Family medicine requires effective dissemination of its growing research base to inform practice, education, and policy. The new Consensus Reporting Items for Studies in Primary Care (CRISP) guidelines may contribute to success. We will describe the pathways primary care (PC) research follows from presentation to publication and test if encouragement to use the CRISP guidelines is associated with increased acceptance and publication rates. We are conducting a confidential online survey in two phases of everyone who presented original research at the November 2023 meeting of NAPCRG (North American Primary Care Research Group), using the Qualtrics platform. Currently in progress, Phase 1 collects data on presenters, studies, research reports, author teams, submission processes, acceptance rates, and publication outcomes. Bivariate and multivariate analyses will identify factors associated with submission, acceptance, and publication. In Phase 2, a randomized controlled trial (RCT) will assign participants to either an observation-only control group or an intervention group receiving the CRISP guidelines. A follow-up survey at 6-9 months will assess presenters’ experiences with acceptance and publication of their written reports. The ongoing Phase 1 survey of 659 presenters worldwide includes diverse professions, specialties, scientific disciplines, and research roles. The presented studies include a broad range of research methods, study designs, problems, populations, and research questions. We will describe presenter experience with the submission, acceptance, and publication of these studies and examine associations with characteristics of the researchers, studies, and reports. The later Phase 2 RCT and follow-up survey will test for differences between the CRISP guideline group and the observation-only control group in success with acceptance and publication. Study results will describe the current practices and patterns of submitting and publishing reports of PC research to guide the dissemination and implementation of research findings to help improve patient care and population health.

Early-Stage Alzheimer’s Disease (ESAD) is characterized by the development of beta-amyloid aggregates (Aβ42) and phosphorylated tau (pTau) leading to mild cognitive decline and variable personality changes. Because specific diagnostic criteria have not yet been established for ESAD at middle age, there is no way of knowing who might be susceptible and who might be resilient to more severe neuropathology and dementia in later years. The geroscience concept assumes pathways associated with aging are also associated with age-related diseases including ESAD. Therefore, a simple skin biopsy procedure shown to predict resilience to aging in middle-aged mice should be able to predict resilience to ESAD in middle-aged mice. An adeno-associated-viral (AAV) vector system carrying pathogenic components of AD, Aβ42, and pTau, was used to induce ESAD in 23-month-old C57BL/6 mice. Before receiving the AAV vector, 2 mm ear punch biopsies were performed, and the rate of closure was measured over 3 weeks. The study ended when mice were 26 months of age, and the closure rate for each mouse was calculated and correlated with behavioral and neuropathological features of EASD. Preliminary observations will help address the question of whether the healing rate of a simple skin wound can predict susceptibility to the burden of AAV-mediated ESAD. It is expected increases in physical resilience will be associated with increased wound closure, and thus, mice with increased wound closure will have greater resilience to the onset of ESAD neuropathology. This could have highly impactful implications for the early treatment of ESAD in human patients thus preventing the irreversible and fatal progression of dementia associated with late-stage AD. In addition, DNA from skin biopsy cores could be used to obtain DNA methylation signatures for determining biological age thus providing an enriched, translationally relevant data set.
 

Individuals exposed to chronic, severe, and/or inescapable stress may adopt dissociation as a coping mechanism to alleviate suffering. In dissociation, individuals report losing connection to their bodies, reality, memories, etc. Dissociative experiences are hallmark symptoms of psychiatric disorders such as Post-Traumatic Stress Disorders (PTSD) and Dissociative Identity Disorder (DID), which has high comorbidity with personality disorders. Maladaptive personality patterns can be thought of as chronic patterns of behavior that are resistant to change. Brain systems are known to underlie these sorts of habitual learning. The interactions between dissociative experiences and action-learning mechanisms are still poorly understood. In this project, we review the history of dissociation and action-learning research and synthesize a model of how dissociative mechanisms may interact with action-learning mechanisms to develop pathological behavioral patterns and even multiple personalities. We propose employing the PRISMA method to select relevant literature from PubMed or other medical databases systematically and conduct a comprehensive review across the spectrum of dissociative disorders. By utilizing the Dissociative Experiences Scale (DES) scores as a measure method, we aim to identify action-learning brain region activity (BOLD signal) and volume changes and explore how these changes correlate with dissociative symptoms; and we’ve found that DID, Dissociative Disorders, and PTSD has the highest DES mean scores. Beyond this, our findings show that in these patients with different kinds of dissociative disorders, for the patients with different kinds of dissociative disorders, their Hippocampus and Amygdala are smaller while their Pallidum Volumes in their brains become typically larger. Despite this relationship having elementary processes, combining with future findings and discoveries, the relationships between dissociative experiences and action-learning mechanisms will be better understood.

The composition of metropolitan governance has many effects on land use decisions, budget allocations, housing development, transportation planning, and racial, economic, and social equity in urban areas. However, there has been little academic inquiry into the effect of regional governance structure on transportation accessibility. This paper seeks to examine statistical linkages between regional governance fragmentation and trends toward and away from greater transportation accessibility in metropolitan areas. I perform a comparative statistical analysis of 47 of the 50 largest Metropolitan Statistical Areas, examining census data from 2002 to 2022 and transit accessibility data from the University of Minnesota Accessibility Observatory from 2014 to 2021 to examine this relationship. The causal factor I investigate is metropolitan governance fragmentation, which I capture through a Governance Fragmentation Index (GFI). The dependent variable, transportation accessibility, is captured through an Accessibility Gap Index, which categorizes transportation access through accessibility levels throughout each Metropolitan Statistical Area, utilizing data from the Accessibility Observatory from 2014 to 2021. My analysis controls for potential confounding variables, such as geographic area, population size, poverty levels, and region. I expect to find that lower levels of governance fragmentation in a Metropolitan Statistical Area will be associated with greater gains in transportation accessibility. Whether or not a significant relationship is identified, the research conducted will contribute to literature and ongoing research surrounding metropolitan governance and transportation accessibility.

Every year, substance abuse, mental disorders, and psychotic behavior become more and more severe, especially in the local communities of Seattle. Our research explores the interconnection between psychology and the utilization of modern computer vision and emotion recognition tools to analyze human semantics/behavioral patterns in order to comprehend the underlying dynamics that lead to these behaviors and ultimately, spearhead efforts for prevention. As a computer science researcher, I employ advanced machine learning algorithms built upon open source programs to track and interpret gestures, facial expressions, and body language in video data. By filtering the data through a database and comparing emotions through statistical analysis in similar individuals, I am able to uncover nuanced patterns in behavior and emotion that a human researcher would otherwise not notice. While the study is ongoing, our current data illustrates correlations between specific gestures, facial expressions, and emotional states in hallucinogenic use. Though these individuals exhibit typical behavior of substance use, they have different backgrounds and contexts that lead to unique obscure changes in emotion and behavior, and our program allows us to identify and analyze these transformations that current medical knowledge overlooks. These findings hold promising implications for informing future research directions and intervention strategies across multiple domains, from understanding human behavior in social interactions to enhancing human-computer interfaces.

Ethnic/racial minorities often face challenges associated with adjusting to a dominant or new host country. These challenges, known as acculturative stress, include difficulties with behavioral, emotional, and social adaptations, and are linked to adverse outcomes. Understanding the degree to which, for whom, and in what contexts acculturative stress may affect mental health is important in designing culturally-informed interventions. Yet, there are several limitations in the current literature on acculturative stress. First, most studies focus on Latino or Asian ethnic groups or international students/cross-national migrants; it is unclear how acculturative stress is differentially associated with health across populations. Second, most studies have focused on mental health outcomes, whereas other important outcomes (academic, relationship) are neglected. Using records from a large-scale meta-analysis project, we conducted a scoping review of research on acculturative stress to characterize the heterogeneity across studies. We performed literature searches using keywords (e.g., “acculturative stress,” “bicultural stress”) in databases including PsycINFO, and identified 3746 relevant studies. Abstract and full-text screening yielded 681 published and unpublished articles eligible for quantitative analysis. Primary studies were included if they measured acculturation in the context of intercultural adaptation and migration, acculturative stress, and health, academic, and relationship outcomes. We coded and summarized sample characteristics of all articles (e.g., % immigrants, % female/woman, mean age). We will randomly select 50 articles that examine acculturative stress and its associations with mental health, academic, and/or relationship outcomes. We will review and present common outcome measures, instruments assessing acculturative stress, and conclusions. We expect the scoping review to indicate patterns, variability, and gaps in the acculturative stress literature. Results will inform future research on overlooked outcomes and understudied populations, and shed light on necessary basic scientific information (e.g., mediators, modifying conditions) that will support the development of culturally-informed interventions and policies.

Advances in language models have made diverse applications in social and health sciences possible. With many models emerging daily, it is important to understand general principles behind how language models represent text data. The study aims to investigate the effects of text manipulation on the numerical representation of meaning, or embeddings, through language models. The research is focused on numerous text manipulations (ex. changing direction of sentiment, sentence incompleteness, object of reference, intensity of statement). The research employs SentenceTransformer for embedding generation and UMAP for the visualization of semantic shifts. A comprehensive analysis of over 50 sentence manipulations provides a preliminary basis for understanding the nuances of semantic space organization. The core hypothesis suggests that incomplete and intensity based modifications lead to greater shifts compared to negative and referencing modifications. This outcome is expected to be substantiated through measures of embedding displacement from original text, and generalized through statistical validation, with tests accounting for multiple comparisons. This research is positioned to scale towards publishable results, aiming to systematically quantify the effects of semantic modifications across a larger dataset. The anticipated contributions of this work are twofold. It offers insights into the organization of semantic space as influenced by sentence modification, providing implications for the development of more nuanced NLP (Natural Language Processing) applications. And crucially, it lays a foundational basis for understanding how large language models (LLMs) process psychological statements and terms, an area of increasing importance as these models become more integrated into applications involving psychological analysis. Overall, the outcomes of this research are poised to deepen the understanding of how semantic alterations impact sentence embeddings, with potential applications in enhancing NLP models' interpretability in processing complex linguistic constructs.

Individuals exposed to chronic, severe, and/or inescapable stress may adopt dissociation as a coping mechanism to alleviate suffering. In dissociation, individuals report losing connection to their bodies, reality, memories, and more. Dissociative experiences are hallmark symptoms of psychiatric disorders such as Post-Traumatic Stress Disorders (PTSD) and Dissociative Identity Disorder (DID), which have a high comorbidity with personality disorders. Maladaptive personality patterns can be thought of as chronic patterns of behavior that are resistant to change, which often are heavily present in brain systems that focus on habitual learning. However, the interactions between dissociative experiences and action-learning mechanisms are still poorly understood. In this project, we review the history of dissociation and action-learning research to synthesize a model of how dissociative mechanisms may interact with action-learning mechanisms as a cause for the development of pathological behavioral patterns and even multiple personalities. We propose employing the PRISMA method to select relevant literature from PubMed or other medical databases systematically and conduct a comprehensive review across the spectrum of dissociative disorders. By utilizing the Dissociative Experiences Scale (DES) scores as a measure method, we aim to identify action-learning brain region changes and explore how these changes correlate with dissociative symptoms; thus far, we’ve found that DID, dissociative disorders, and PTSD have the highest DES mean scores. Beyond this, our findings show that in these patients with different kinds of dissociative disorders, the hippocampus and amygdala are smaller and the volume of the palladium is typically larger. When combined with future findings and discoveries, the relationships between dissociative experiences and action-learning mechanisms will be better understood.

Most research on health and food choice is conducted with samples of college students with low racial and socioeconomic (SES) diversity. The University of Washington, Tacoma (UWT) has a uniquely heterogeneous student body, composed of various ages, SES, and races, but the aspect of non-traditionality that interested us was our large population of commuting students. The economic hypothesis for overnutrition highlights the health disparity created by the inverse relationship between food price and diet quality. Yet, in this population, we do not see a significant difference in body composition across income (F (3, 315) = 1.137, p = 0.334). However, there is a significant negative correlation between perceived stress and household income (r = -0.119, p < 0.05). Typically, time scarcity drives convenient food choices and consumption of sugar-sweetened beverages (SSB), often associated with adverse health indicators. In this project, I asked if longer commute times (zip codes farther away from campus) contribute to economic choices (money spent on gas, less for food) and time scarcity. I hypothesize that individuals with zip codes farther from campus (longer commute time) will show greater implicit preferences for SSBs and be more likely to select SSBs in a laboratory study. An independent sample t-test showed no significant relationship between commute time and SSB selection, t (318) = 0.875, p = 0.382. Again, a chi-square test showed no significant association between SSB selection and SES category 𝝌² (4, n = 318) = 2.505, p = 0.644. There is a modest inverse correlation between SES and implicit wanting for SSBs, r = -0.099, n = 291, p < 0.10. Addressing SSB consumption from a unique perspective accounting for external and personal factors with diverse sample populations is crucial to addressing the personal responsibility narrative in the psychology of eating.

Humans have a tendency to display greater empathetic and prosocial responses to in-group, compared to out-group members. Within the context of psychological treatment, learning from others such as a therapist or a support person about what is safe or dangerous is an important skill for therapeutic success, yet studies have not systematically examined how group membership based on gender impacts social fear and safety learning. Our study aims to investigate the impact of having the same gender as the learning model on learning safety from others. We will assess extinction learning by measuring skin conductance responses to safety cues versus vicarious reinforcement cues during a vicarious extinction learning task. This project builds upon previous studies showing enhanced safety learning between individuals belonging to the same social groups and aims to examine the interaction between gender categorization and social anxiety. 100 participants (50 Social Anxiety Disorder (SAD), 50 Healthy Control (HC)) will be recruited to undergo an extinction task based on an established fear learning paradigm featuring a male learning model. The effect of gender matching on vicarious extinction will be examined by comparing responses between non-male-identifying (out-group) and male-identifying (in-group) participants. We hypothesize that those with the same in-group gender categorization as the model (Gender-Matched) will benefit more from vicarious extinction, as compared to out-group members (Gender-Mismatched). This is evidenced by lower skin conductance responses to learned safety cues during the Extinction and Reinstatement phases of the task, and we anticipate that this effect will be stronger for those in the SAD group. Findings will highlight a potentially significant moderator of vicarious fear extinction and may provide a mechanistic explanation for enhancing fear extinction recall for patients with SAD.

Family businesses are some of the most important economic contributors in the United States, accounting for approximately 64% of the U.S. GDP. The family business model, which refers to any business with two or more family members on the board or in ownership, is a crucial and enduring part of business in the Seattle area and abroad. Historians have often pointed out that the family business model seems to be the base model for business and has thus been present since the beginning of organized business, often in the form of farms, merchant companies, banks, and other small businesses. Despite its prevalence, the family business model is far from perfect because of its numerous commonly encountered limitations. One of the limitations family businesses face is the challenge of succession, as only about 30% are able to succeed from the first generation to the second. Other limitations relate to growth, sustainability, and qualification problems. This study, conducted as a literature review, uses a combination of peer-reviewed articles and popular sources (chosen based on criteria of relevancy and prominence) as quantitative data to examine the consensus of family businesses in Seattle and the solutions that have been proposed to address these limitations. Interviews with family business owners in the Seattle area were also conducted to provide qualitative data and to highlight specific opinions. The economic and historical implications of Seattle family business are also discussed. This research aims to provide insight into otherwise costly financial, succession, and leadership difficulties in order to ensure that the family business model is an enduring contributor to the Seattle economy. Having the proper knowledge on how to approach these difficulties and reconcile with their seemingly conflicting nature can help family businesses in the Seattle area thrive while working through complicated business situations.

This research project explores the relationship between community social capital and corporate gender diversity, focusing on female executives within S&P 1,500 firms. Previous studies have shown that it is important to have more women in top leadership roles in business organizations today because it provides different perspectives and facilitates various leadership styles, promoting innovation and driving performance. Unfortunately, during the sample period of 7 years between 2010 and 2017, only 8% of top executives in C-suit at S&P 1,500 firms were female. To understand why this is the case and suggest a way to improve this case of female underrepresentation in executive positions, I study the community social capital that fosters a culture of trust, cooperation, and mutual respect, which are foundational for creating inclusive work environments that value diversity. I hypothesize that substantial regional social capital promotes a higher representation of women in top management roles. I collected county-level social capital data from Penn State’s Northeast Regional Center of Rural Development (NRCRD) and gender diversity data from the WRDS ExecuComp database. To test my hypothesis, I conducted a regression analysis. Empirical results indicate that sample firms with greater exposure to higher community social capital are more likely to appoint women to top management positions, which allows them to make crucial decisions within the organization. This study highlights the impact of community dynamics on corporate policies and displays a new perspective on promoting gender diversity in leadership positions. This project intends to improve an individual learning experience by providing valuable insights into the interconnection between societal structures and corporate governance, concentrating on gender inclusivity.

Alcohol and other substance use are associated with a range of negative physical, mental, and social consequences including blackouts, unintended injuries, involvement in interpersonal violence, and other health problems. Young adults in college are particularly at risk for alcohol and other substance use. While existing research on reduction interventions often emphasized individual responsibilities of those that engage in alcohol and substance use, peer interventions remain an overlooked approach to harm reduction. Bystander interventions have been shown to be effective in preventing and reducing the harm of sexual assaults among college students. However, there is less empirical attention on applying bystander interventions to reduce the negative consequences of substance use. An important first step of understanding how to incorporate bystander interventions to address substance use is to have a reliable and valid measure assessing these behaviors. This study aims to develop and examine the psychometric properties of a new bystander behavior scale specific to substance use. Data came from a large multi-site survey study with 930 college students (Mage = 19.39, 69.6% female, 58.1% White). I will conduct exploratory factor analysis to extract underlying factors that best explain the observed correlations between the items. Next, I will examine convergent and discriminant validity of the scale scores by correlating them with established instruments such as Drinking Refusal Self-Efficacy Questionnaire and Personal Assessment of Responsible Drinker Identity. Individuals who score higher on the bystander behaviors measure are expected to score higher on self-efficacy in resisting alcohol and be more likely to identify as responsible drinkers. Results will provide initial validation data for the bystander behaviors measure and can be used in future research and intervention implementation that address substance use among college students.

Air pollution is a key component to understanding the Public Health of populations globally, with Diesel Exhaust Particles (DEP) being a significant contributor to traffic-related air pollution. Exposure to DEPs varies across populations and is therefore crucial to understanding the continual impacts of traffic-related air pollution on the public. Prior research has indicated that the formation of Amyloid-𝛽 (A-𝛽) plaques and activation of the  nucleotide-binding domain, leucine-rich–containing family, pyrin domain–containing-3 (NLRP3) inflammasome is linked with the development of Alzheimer’s disease (AD) later in life. AD is a form of progressive disease that impairs memory and other cognitive functions and impacts the lives of tens of millions of people globally. This study aims to confirm the linkage between exposure to DEP and memory impairment through NLRP3 inflammasome activation, utilizing an animal model to investigate a potential increase in AD later in life. We exposed male and female low-density lipoprotein receptor knockout (LDLR KO) mice chronically to inhaled DEP or filtered air as a control for 18 weeks. We then utilized the Object Location Memory (OLM) and Object Recognition Memory (ORM) behavioral tests to investigate the immediate impact of multi-week DEP exposure on short-term memory, another indicator in AD progression. Afterward, we sacrificed the mice and harvested a variety of tissues, including the brain. I conducted Immunohistochemistry (IHC) on cryosections of the exposed and non-exposed brain to assess DEP-induced AD-like brain architectural changes and to quantify the impact of DEP exposure in activating the NLRP3 inflammasome, ultimately leading to neurotoxicity, and to the development and progression of AD. Confirming the association between diesel exhaust and the NLRP3 pathway provides a potential therapeutic target in populations at an elevated risk for AD.

Advances in language models have opened up endless applications in the social and health sciences. A key to fulfilling this promise is the development of rigorous practices for gathering data used for model training. This research project reports the development of standardized protocols for building databases for self-reported psychological questionnaires that can be used to fine-tune language models. These questionnaires cover a spectrum from childhood experience to altered states of consciousness and beyond. We systematically review the literature on appropriate database-building strategies, use combined data collection techniques, and instill specific criteria into our collection process to help facilitate the research’s computational pipelines on evaluating the relationship between semantics and psychological factor membership. Using a 2D semantic map of the PsycTests database as a reference, we can further evaluate the effectiveness of our protocols by developing quantitative measures to assess comprehensiveness and objectivity in our database building. Our initial explorations involve measuring data variance and density of data distribution. This research provides a general toolkit for building text databases for appropriate and effective model training purposes across various disciplines.

On February 24, 2022, Russia began its full-scale invasion of Ukraine. Washington state was already the home of over 42,000 Ukrainian immigrants, and since the invasion, upwards of 22,000 Ukrainian refugees have arrived. Refugees face significant challenges due to obstacles stemming from the trauma of war, displacement, resettlement, and the lack of access to essential health and social services. The Nashi Immigrants Health Board is a non-profit, community-based organization founded by public health professionals. Nashi partners with the Ukrainian community via health promotion, education, and advocacy programs and provides culturally and linguistically appropriate public and mental health services. Nashi’s warm-line was created as a caller line for Ukrainian-speaking communities, primarily refugees fleeing Ukraine to provide psychosocial support and connection to community resources. The purpose of this study is to examine the data collected during warm-line calls to identify the greatest needs and streamline resources for the community. Warm-line operators collect anonymous information such as age, zip code, primary language, the purpose of the call, and action taken are recorded. Caller data are entered into a database with specific categories and analyzed for trends and patterns of needs, which will help Nashi refine the pathways to interventions and resource allocation. We hypothesize the callers will be mainly calling about needs such as housing, health insurance, and job opportunities. We expect the navigators to provide emotional support in addition to resources, even if callers are not expecting or requesting it. The findings would help Nashi inform the Office of Refugee and Immigrant Assistance (ORIA) for future programming and quality improvement interventions. The study results are not inclusive of every need in the Ukrainian refugee population. Ongoing efforts of resource allocation would continue to evolve based on the changing needs of the Ukrainian community.

The potential for pathogenic disgust to augment bias is clear in the wake of the COVID-19 pandemic. Yet, existing research overlooks the specific link between the experience of disgust and expression of weight bias. In contrast to the typical research focus on overnutrition from the perspective of individual weight management, I examine how internalized weight stigma – among other factors – could moderate the relationship between types of disgust sensitivity and bias against people with a larger body size. My hypothesis is that increased disgust sensitivity will predict weight bias. Specifically, I expect this relationship between pathogenic disgust and weight bias to be moderated by cognitive rigidity, internalized weight stigma, and interoceptive accuracy. In my online study, undergraduate psychology student participants complete the 32-item Disgust Scale – a measure using a verbal paradigm for disgust. My other validated psychological instruments included the Detail and Flexibility Questionnaire, the Interoceptive Accuracy Scale, a modified version of the Weight Self-Stigma Questionnaire, the Fat Scale from the Universal Measure of Bias, and the Daily Inventory of Stressful Events. My goal is to discover the nuanced relationships that govern the manifestation of weight bias. The outcome of my study has implications in predicting contributors to weight stigma and thus designing interventions to prevent weight bias in the healthcare profession, the general population, and the field of food psychology itself.

Individual eating behaviors, and the motivations behind food choices, impact a person’s overall health and wellbeing. While cognition plays a significant role in food decision-making, processes outside of conscious awareness like high levels of perfectionism (cognitive rigidity) can influence a person's eating habits. On the other hand, these choices are impacted by emotional processes, as a person’s interoception sensitivity levels and experience of interpersonal stress impact food intake. The notions of cognitive rigidity and interpersonal stress contributing to changes in eating habits are contrary to the current discourse of personal accountability surrounding food choice and diet. The personal accountability narrative emphasizes each person’s individual responsibility in deciding what to consume, the quantity, and quality of the food. In contrast to the idea that each person is responsible for their health, analysis of surrounding factors that contribute to the decision-making process yields a more accurate understanding of health and well-being. Therefore, I aim to explore the relationship between interoceptive processes, interpersonal stress, and cognitive rigidity to explain food decision-making and diet quality. To examine this relationship, I am recruiting a sample of college students from psychology courses at the University of Washington Tacoma campus. Participants will complete an online self-report questionnaire assessing interoception, perfectionism, mood, interpersonal stress levels, and eating habits. I hypothesize that interpersonal stress and cognitive rigidity adversely impact interoceptive sensitivity, ultimately worsening participants' diet quality. Understanding the relationship between interoceptive sensitivity, interpersonal stress, and cognitive rigidity and their impact on well-being can help inform future health interventions guiding improving dietary consumption.

Over the past several years, the city council of Oxford, England has developed an environmental policy project named the Zero Emissions Zone (ZEZ) which seeks to establish an area in the city center where vehicles will be either prohibited or fined for through travel. The council conducted many studies on where the policy would most effectively reduce carbon emissions from private vehicle use and considered existing pedestrian-only areas for potential use. The policy has proven controversial in the community. There has not been significant research on the popularity of the policy and how environmental policies like the ZEZ affect the political beliefs of Oxford community members. I hypothesized that the ZEZ would be a popular policy on the whole with the primary opposition coming from those who utilize private vehicles as their primary mode of transportation in Oxford and that approval or disapproval of this policy will significantly affect one’s perceptions of the city council and political parties. Through an anonymous online survey, I collected data from respondents over the age of 18 who either work, live, or study in Oxford or the surrounding suburbs. Participants answered questions regarding their political opinions, views on the ZEZ policy, and how the policy has affected their politics. These questions sought to illuminate a connection between views on environmental planning policies and political perceptions in general. I expect the results to show that younger respondents, particularly those whose political beliefs are to the left of center, will approve of this policy, and by extension the city council, more so than older respondents. This research has implications for Oxford City Council and any others interested or intending to install or expand similar zones so that they may understand what impact it may have on local politics. 

In recent years, organizations have debated whether elements of “white supremacy culture”, including a sense of urgency, may create inequitable environments for people of color. We aimed to empirically investigate whether cultures of urgency in professional settings undermine the recruitment of racially minoritized groups. Undergraduate participants (N = 219) read job advertisements for two jobs: one where urgency culture is highly valued (e.g., “swiftly reset priorities at any given time”) and one with less sense of urgency (e.g., “adjust priorities based on capability”; order counterbalanced). Participants then reported how likely they would be to apply to each job. We hypothesized that individuals from racially minoritized groups will be more likely to apply to lower-urgency jobs over high-urgency jobs. The results show that most participants preferred lower-urgency jobs. However, countering our hypothesis, individuals from racially minoritized groups held more positive attitudes towards high-urgency jobs (e.g., how well they think they'd perform in the job) when compared to white participants. This finding casts doubt on the proposed elements of "white supremacy culture". It urges for more empirical research on how different racially minoritized groups may perceive these elements in work settings. Furthermore, our sample consists mainly of Asian Americans, which does not speak for the experiences of other racially minoritized groups. Our future research will focus on diversifying samples collected.

The COVID-19 pandemic spotlighted Asian American (AA) mental health and their foreign position, underscored by anti-Chinese rhetoric and policies and the concurrent surge in hate incidents, crimes and xenophobia against AAs. However, framing COVID-19 as the sudden cause of anti-Asian hate facilitates misconceptions about AAs: it undermines a storied history in which AAs have long survived and challenged their treatment as “forever foreigners.” To understand the extent to which anti-Asian hate and mental health challenges among AAs during COVID-19 was a unique event in AA racialization, it is paramount to understand AA marginalization and its implications for mental health prior to COVID-19. To do so, I use the 2016 Collaborative Multiracial Postelection Survey (CMPS). The survey gathers data on respondent attitudes across a diverse set of political, psychological and socio-demographic measures and is ideal for Asian ethnic subgroups and intersectional analyses of gender, age and immigration generation. My analysis includes bivariate and multivariate regression analyses using the sample of AAs (N= 3006). Thus far, I have found that a substantial proportion of AAs indicate low feelings of belonging in the U.S., personal experiences of racial discrimination, and depressive symptoms. For the project’s next step, I will examine the extent to which AAs share a common foreign experience prior to COVID-19 and how this contributes to depressive symptoms. I expect that there will be significant subgroup differences at the intersections of ethnicity, gender, and immigration generation in the relationship between experiences of racial discrimination, feelings of belonging, and depressive symptoms when community involvement and local contextual factors are considered. This work helps situate anti-Asian hate during the pandemic as part of a more extensive history of racism in the U.S. and broadly, contributes to more informed mental health services by highlighting the implications of racism for mental health.

Disparities and consequences arise for pregnant individuals struggling with substance use disorder. Policies aimed at preventing substance use during pregnancy cause harm instead of providing support and purposeful treatment options. These policies that are deeply rooted in systematic racism have been shown to disproportionately affect the Black, Indigenous, and people of color communities, leading to increases in adverse birth outcomes. In 2020, approximately 8% to 11% of pregnant individuals between the ages of 15 to 44 reported using illicit substances, tobacco, or alcohol during their pregnancy. Addressing the pressing need for improved prenatal and postpartum support for these individuals is imperative. The purpose of this study was to focus on the upstream prevention of neonatal abstinence syndrome, with the overall aim of creating an agenda outlining viable interventions recommended by five specialized healthcare professionals. We conducted extensive interviews with healthcare experts researching prevention policies and those working directly with pregnant individuals struggling with substance use disorder. Additionally, we compiled data through a systematic review of both punitive and supportive policies regarding substance use during pregnancy. We hypothesized that punitive policies are adversely affecting pregnant individuals by creating disparities and hindering proper pregnancy care. The study findings will allow us to design viable interventions aimed at improving pregnancy care for these individuals and ultimately prevent neonatal abstinence syndrome.

Music interventions are gaining attention due to the evidence they improve health and cognition in elders. In the US, there is an increasing number of Mandarin-speaking elders whose cultural background affects their aging experience. After English and Spanish, Chinese is the third most spoken language in the US. Three and a half million households in the U.S speak Mandarin or Cantonese. However, there is little information regarding specific music choices or culturally-tailored music interventions for Mandarin-speaking elders to improve their health and aging experience. The purpose of this study was to gain insights into Mandarin-speaking elders’ views of aging, ageism, and the relationship between music and health. We conducted a qualitative study that involved interviewing Mandarin-speaking elders in Mandarin. The interview consisted of questions related to their cultural perspectives on aging, experience with ageism in the US, and the impact of music on their health. The participants selected music to play during the interview and discussed what the music meant to them and their thoughts on how music affects their health. Four interviews in Mandarin have been conducted to date. Initial findings reveal that filial piety and respect are recurring cultural themes that influence participants’ perspective and experience of aging. Internal ageism was identified in one of the interviews while the participant discussed their experience with ageism. Participants expressed that music benefits their health by providing emotional support and outlet, motivation, reminiscence of the past, brain activity, and stimulation through physical movement such as dance. The findings increase our understanding about the role of culture in the aging experience and music’s role in enhancing health for Mandarin-speaking elders. Future studies can use this information to develop music-based interventions tailored for Mandarin-speaking elders to improve their health.

Approaches to learning new languages and cultures, among other subjects, are ever-changing alongside advancements in technology. In this study, I examined the informal self-regulated English as a foreign language (L2) and intercultural learning of a native Japanese (L1) speaker through their interactions with social media platforms such as YouTube, Discord, and X (formerly known as Twitter). In an online semi-structured retrospective interview with "Yuki," a Japanese woman in her thirties, I queried three areas of informal online language and intercultural learning: 1) her language learning background and the various media she interacts with; 2) her perspective and attitude towards self-regulated learning, her motivations, and how this compares to formal learning; and 3) how her connections to multiple communities on various platforms have influenced her learning and outlook on life. This interview was conducted in Japanese, to allow Yuki to describe her thoughts and experiences as clearly as possible. Findings show the strong impact of online communities on Yuki's motivations and objectives. The global community she encountered offered plentiful opportunities to expand her outlook on life, from her views on pronunciation to the importance of identities. Through the internet, Yuki found others with similar learning goals and began hosting weekly online book club sessions, where individuals of varying English levels are able to develop skills in collaboration with one another despite never having met in person. Along with the possibility for self-directed informal learning, other forms of online content, such as podcasts, allow Yuki to freely teach and challenge herself to interact with everyday spoken English, such as slang and diverse dialects. Although this study only consists of one participant, findings show the tremendous developmental potential of online spaces for language learners. 

Compassion fatigue, also known as secondary traumatic stress or vicarious trauma, is a condition characterized by emotional and physical exhaustion, often experienced by individuals who provide care, support, or assistance to people who are suffering or traumatized. This phenomenon commonly affects professionals in helping and caregiving roles such as healthcare workers, social workers, counselors, and first responders as they’re constantly exposed to secondary traumas. This study investigates five domains of worker well-being of social services employees at a local Homeless Services Agency in King County. Seventy-five social workers and case managers were invited to participate in the online, anonymous survey that contains the National Institute for Occupational Safety and Health (NIOSH) Worker Well-being Questionnaire. This questionnaire consists of sixty-eight questions related to work evaluation and experience, workplace policies and culture, workplace physical environment and safety climate, health status, and home, community, and society. We hypothesize that those who have been employed at the Compass Housing Alliance for more than 2 years will exhibit higher compassion fatigue through their survey responses. The insights gathered will aid in understanding the working environment on service employees’ well-being. Results will offer areas to support workers and hopefully serve to mitigate the potential development of compassion fatigue among staff members.

As the percentage of individuals facing homelessness rises, many organizations working closely with unhoused populations aim to secure consistent housing and essential resources for those who lack this necessity. There are roughly 14,000 individuals facing homelessness in the greater Seattle area. Previous studies have shown positive correlations between employee well-being and job performance. Our research aims to investigate worker well-being among staff members who are currently employed at the Compass Housing Alliance, an organization working closely with unhoused populations to provide both housing and access to other essential needs. This research utilizes the National Institute for Occupational Safety and Health (NIOSH) worker well-being survey. We collected data on five factors that influence the perceptions of 75 staff members on their overall well-being including workplace safety, mental wellness, leadership support, coping strategies and job satisfaction. Expected results from our survey indicate that staff members' perceptions significantly impact their overall well-being. Collecting data using quantitative methods enables us to gain a deeper understanding of these current perceptions in order to utilize relevant resource implementation in areas where gaps are identified to support the needs of staff and improve their well-being.

Including undergraduate peer facilitators (UPFs) in lectures can support students in large and often challenging introductory science courses. Our team decided to increase UPF involvement by creating BIOL113, an undergraduate-led companion course to BIOL180, the largest introductory Biology course at the University of Washington. This study is part of a larger project that assessed the overall experience of both UPFs and students, and this particular work focuses on the former. I investigated how BIOL113 contributed to the experience and development of UPFs.This study incorporated qualitative data from graduated and enrolled UPFs alongside UPF applicants. Through surveys, I explored personal, professional, and academic growth, leadership skills, program expectations, and a sense of belonging. My findings underscore the distinctive role of undergraduate-led programs in enhancing UPF experiences. UPFs’ expectations for the program aligned with their experience and enriched their sense of belonging within the biology department, STEM, and the broader University of Washington community. Furthermore, engaging in UPF roles developed individual leadership skills while also contributing to their personal, professional, and academic development. In addition, the survey highlighted an important area for improvement: tailored professional development for UPFs should focus on marketing the skills gained in teaching and leadership as particularly valuable in the job market, graduate school, or professional school. Understanding the impact of BIOL113 on UPF experience can inform instructional approaches, with the possibility of expanding such initiatives university-wide.

Evidence-based changes to instruction can lead to better student outcomes and performance. For this reason, instructors are interested in collecting data about student experience and outcomes. However, the process of data collection and analysis can be time-intensive for instructors, making it challenging to gather data necessary to improve their classes. Moreover, when data collection does happen, it often centers quantitative data, but this systematically devalues students' experiences. Therefore, we sought to develop a tool that aids instructors in processing and analyzing qualitative data pertaining to students’ class experiences. As part of our research project, we have developed an R-shiny based data processing app that integrates Artificial Intelligence to summarize findings from open response questions on student surveys. This tool is intended to alleviate the time-intensive nature of analyzing student responses to open-ended survey questions. In order to validate the accuracy of AI-generated summaries, we compared them to manual summaries generated using in vivo qualitative coding methods. We find that our app generates responses that are comparable to the manually generated summaries. The summaries include prominent themes along with details about student experiences within those themes. With this tool, instructors can gather real-time data, even in large classes, eliminating the concern of the time-intensive process of manually reviewing responses. By developing this tool, we hope to empower instructors to explore diverse questions that provide them with valuable insights on how to optimize the structure of their classes to improve student experience and outcomes.

 Music is considered a therapeutic tool to improve physical, cognitive, and emotional health. The nature of music varies across cultures and backgrounds around the world. This research project explored the connections between music, aging, and ageism in older adults from Kenya and the therapeutic role of music in improving physical, cognitive, and emotional well-being. This project considers cultural similarities and differences between Kenya and the United States and examines the unique nature shaped by different cultures. In this study, we explore how music affects the perception of age in older adults from Kenya and their perceptions of ageism. This research project focuses on using interpretive research method where data is collected through interviews to gain a deeper understanding of the topic. The study involves three older adults from Kenya between the ages of 60 and 65. The interview questions focus on understanding the cultural influences of Swahili speakers in the context of music and ageism. Through research on the differences in aging between Kenyan and American older adults, this project seeks to identify cultural factors and explore ways that music can enhance well-being among older adults. This project provides valuable insight into cultural aspects that influence aging and the potential benefits of music-based interventions. The findings have the potential to improve our understanding of how cultural differences influence attitudes toward aging and the impact of music on the health and well-being of older adults.

As healthcare advances, older people with HIV (PWH) have a longer life expectancy. Biotechnological advancements, such as antiretroviral therapy (ART), suppress HIV replication, resulting in decreased mortality among PWH, while increasing quality of life. However, despite these advances, older PWH generally experience an earlier onset of comorbidities due to increasing age, increased adiposity related to HIV medications and sedentary lifestyle, poorer physical function, and high symptom burden. Physical activity is a significant non-pharmacological approach to mitigate chronic disease in older adults and promote healthy living. Among different physical activity types, exercise can be defined as structured and repetitive bodily movement, objectively improving or maintaining physical function. Exercise can take many forms and be performed at different intensities. HEALTH Study participants perform continuous moderate exercise (CME) of walking or jogging at 50% of heart rate (HR) or high-intensity interval training (HIIT) of alternating short intervals of high- and low-intensity aerobic exercise at 80-90% and 50% HRmax. Healthy aging among PWH includes decreased fatigue, greater functional mobility, and fewer symptoms of HIV and its related treatments. What is unknown, and currently investigated in the HEALTH Study, are exercise types to improve physical health outcomes in older PWH. This study aims to explore the relationship and associations between exercise interventions on body composition across older PWH participants. In this year-long undergraduate research project, we conducted a secondary data analysis to compare the effectiveness of exercise interventions, HIIT versus CME, on DEXA scan outcomes of body fat and bone density in older PWH. DEXA scans are medical imaging x-ray tests to quantify body composition. We anticipate that with increased physical activity, especially in HIIT participants, body fat will decrease and bone density will increase. These findings may indicate how exercise interventions can mitigate fatigue and provide recommendations to prompt healthy aging in older PWH.

Millions of people in the United States of America are suffering from substance use disorders (SUD); of particular concern is opioid use disorder (OUD) due to the high risk of overdose and death. This epidemic is a public health crisis that impacts people in every community. OUD is especially harmful to vulnerable populations who face arduous challenges in accessing treatment. People with chronic pain or complex comorbid conditions, housing-unstable communities, and the under/uninsured are all particularly vulnerable to OUD. Safe and effective medical treatments are available to mitigate overdose risk, but accessing them can be bureaucratically challenging for providers to navigate, which results in the relegation of OUD treatment to separate specialty facilities. This fragmentation of care impedes access and decreases provider follow-up. Substance users seeking help often face undue burdens in the form of strict sobriety requirements or incremental prescriptions necessitating frequent clinic visits. Madison Clinic at Harborview Medical Center provides care for people living with HIV and AIDS. This population is particularly vulnerable to medical stigma and bias, job instability, homelessness, chronic pain, and substance use. Using a qualitative descriptive method, we conducted a thematic analysis of semi-structured interviews to explore the thoughts, opinions, and perspectives of providers at the Madison Clinic and people living with HIV or AIDS using substances regarding the integration of medical treatments for substance use into the primary care setting. Interview transcripts were analyzed through a mix of inductive and deductive coding. Ideally, this integration would help remove barriers and alleviate the complexity of coordinating fragmented care. We anticipate co-locating routine care and treatment for opioid use simultaneously may lead to better adherence to medication regimens and increased patient and provider satisfaction with care. This research will influence the creation of a healthcare model for integrating substance use treatment into primary care settings.

Games are engaging for children, and educational games make learning fun. Scholars have conducted research on how games help kids learn, but little is known about the specific ways that children’s mental models of real world concepts change through playing games. This study specifically investigates the process of how children learn STEM principles through exploring real world STEM concepts integrated into games. This ongoing research is being conducted with cohorts of middle schoolers in the King County area. Current games include Dragonwood, a card game with elements of probability, a Dungeons and Dragons campaign structured with elements of physics, and a STEM systems game called Oxygen Not Included. As the participants play these games, we ask them questions about their actions, interpretation of the game space, and why they make certain choices within the games. In doing so, we hope to understand the specific learning processes and parts of the game that change their understanding. Through qualitative analysis we are identifying trends in the students’ responses. We expect to find insight into learning through games and particular components in games that are beneficial for learning STEM concepts. This research has implications for educational game design and delivery. Our findings could impact the use of games in classrooms and provide insight for the development of educational games. This research could further make STEM learning more accessible and engaging for children.

The use of deepfake technology to produce pornographic content (deepfake pornography) is a form of sexual exploitation and violence of those most vulnerable to sexual harassment in our society: women and girls. Deepfake pornography, sometimes called synthetic pornography, refers to the process of manipulating an image by adding sexually explicit content. A 2019 study conducted by Deeptrace Labs found that 96% of deepfake technology is used to create non-consensual pornography targeting women and girls. As technology develops, US privacy laws fail to keep pace, leaving online predators unchecked. While there is no federal regulation on the use of deepfake technology, in the past few years, nine states including California, Texas, and New York have passed laws governing their use. The most substantial of which is California's AB 602, passed in January of 2023, which banned the distribution of pornographic deepfake images and videos made without consent and which may result in the defendant paying up to $150,000 in statutory damages. This literature review examines the rise of deepfake pornography used against women and girls, and the impact it has on its victims. It also compares how regulators in the US and certain other jurisdictions are addressing this issue. This review brings awareness to the growing online predatory activity facilitated through deepfake technology and the need for swift action to address this crisis on a nationwide scale.

Nearly 70% of older adults hospitalized in an intensive care unit (ICU) experience delirium, a risk factor for long-term cognitive impairment that persists beyond discharge. The severity of critical illness, coupled with the 24-hour care provided in the ICU, is associated with significant disruptions to sleep and the circadian rhythm. These circadian rhythm disturbances, which affect up to 80% of ICU patients, may decrease the efficacy and benefits of interventions to improve cognitive function. Few intervention studies have been conducted testing circadian-based approaches to optimize timing of interventions to prevent cognitive decline in older ICU survivors. The study aims are: 1) to assess the feasibility, adherence, tolerability of morning or afternoon sessions of a computerized cognitive training intervention, and 2) to explore the role of individual chronotype on intervention usability and acceptability in older ICU survivors. Participants are randomized to one of three arms: morning computerized cognitive training sessions, afternoon computerized cognitive training sessions, or usual inpatient care. Participants assigned to the intervention groups complete daily 30-minute cognitive training sessions for up to 7 days or until hospital discharge. Wearable sensors monitor circadian rhythm patterns (via continuous body temperature and activity/sleep), and participants answer a questionnaire to determine their individual chronotype (“morning” versus “evening” circadian preference). Upon study completion, participants provide quantitative and/or open-ended feedback via surveys. We hypothesize that participants whose assigned intervention timing aligns most closely with their individual chronotype will demonstrate higher intervention adherence and will report higher tolerability and acceptability. Data collection is ongoing; results will investigate the potential of circadian-based and chronotherapeutic interventions to mitigate cognitive impairment in older ICU survivors. Additional research is needed to develop personalized interventions that integrate individual circadian rhythm and chronotherapy as targets to accelerate cognitive recovery throughout critical illness.

There is currently a lack of tractable data on what was published in England in the early 20th century. However, this information exists in printed volumes of The English Catalogue of Books, which have been digitized through the HathiTrust digital library. The English Catalogue of Books, released in the UK by the trade publication Publishers’ Circular, provides a yearly record of books published from the mid-19th to the early 20th century. Each catalog has been converted to plain text automatically through optical character recognition (OCR). Our aim was to parse this plain text into user-friendly data on books published each year in the UK. However, the OCR-generated text often contained errors and inconsistencies that prevented the effective extraction of data on books listed in the catalogs. Thus, we aimed to gauge the accuracy of existing methods for parsing the catalogs and to tailor processes for gathering data from catalogs published between 1908 to 1922. By writing regular expressions to capture, split, and match the patterns of bibliographic entries, we were able to improve the accuracy of processes for extracting data. Our solutions increased the number of publications for which information was accurately being captured by on average 28%, and we were able to record information on over 21,500 books that previously had not been captured. We also created a summary dataset from the catalogs with information on the overall output of the publishing industry during these years. By analyzing and visualizing this publishing data, we were able to show that fiction was the most frequently published genre during the period. We anticipate this project to be the foundation of more work towards efficiently parsing The English Catalogue of Books in order to offer insights into the British publishing industry in the 19th and 20th centuries.

The recognition of individuals as sufficiently "Darwinian" entities for natural selection has led to the exploration of clades as alternative units of selection to species in the field of philosophy of biology. Previous research has demonstrated that clades meet the criteria of "Darwinian" individuals, intriguing philosophers to explain natural selection at the clade level. This research employs an Abstract Data Type (ADT) approach from computer science, constructing a data structure (DS), and attempts to implement the clades ADT both with and without species. The objective is to scrutinize the inner structure of clade and unearth the interplay between clades and species, a relationship not explicitly implied in their definitions. Preliminary findings affirm the technical feasibility of implementing clades ADT as a clade-tree data structure with species, while the feature of each level of clades remains unclear: (1) Issues of trait identification emerge when assigning properties to organisms within clades; (2) The empirical nature of the data used to construct the data structure representing clades limits holistic demonstration of evolution. On the other hand, a clade-tree data structure without species encounters challenges: (1) Historical organisms within inner stages cannot be distinctly separated without defined species-like boundaries (2) If properties are specified to species-level, the species-free intention of the data structure becomes questionable. The emergence of a dependency on species introduces a compelling reason for reconsidering the ability of clades in explaining natural selection, particularly for those philosophers who oppose species as a unit of selection.

This essay dives into three key stories: the myth of Pygmalion and his statue from Ovid's Latin epic Metamorphoses, the story of Acis, Galatea, and Polyphemus also from Metamorphoses, and Madeline Miller's novella Galatea. The major factor that these stories have in common is the overlying theme of male desire, and the consequences of not being able to attain that desire, typically in the form of control or violence. I analyze the literary choices made by both Ovid and Miller, and how they characterize their male characters in juxtaposition with their female characters. I take these analyses and connect them to the modern phenomenon of the Internet "Incel", which is a colloquial term for "involuntary celibate"— an online community of avid misogynists. My main methodology is utilizing critical sources by feminist classicists such as Donna Zuckerberg in conjunction with research studies done on male-dominated online forums to understand how misogynistic rhetoric is formed out of male desire. My essay focuses on Pygmalion and Polyphemus as prototypical Incels, and how this sort of misogyny and desire for control over women has a long history in popular media, and emphasizes the role that the classics play in modern day sexism.

While survival rates from breast cancer have notably improved, survivors still face long-term health complications due to the side effects of chemotherapy. A significant concern is that patients can develop lasting cardiovascular problems that are exacerbated by physical inactivity during treatment. This study hopes to address this by identifying the optimal fitness intervention for improved cardiorespiratory fitness (CRF) to enhance the cardiovascular health of breast cancer survivors, particularly in those who showed reduced CRF after breast cancer treatment. This randomized control trial enrolled thirty breast cancer survivors into an aerobic exercise, resistance exercise, or control group. Over six-months, the participants performed their designated training, and MRI scans were taken before and after the training program. I am analyzing the depots of fat, including visceral and subcutaneous fat from MRI images of the thigh and abdomen in order to determine the association with cardiorespiratory fitness. Statistical analysis utilized a paired t-test with an intent-to-treat approach to determine the difference in fat levels and to mitigate bias from non-compliance and dropouts. I expect a marginal difference in levels of overall fat between the separate fitness interventions, however, I anticipate a significant difference between the control and both fitness groups. The interpretation of this data provides a first step towards understanding how exercise interventions can be tailored to breast cancer survivors, potentially improving post-treatment survivorship.

Déjà Vu is an emotional experience that occurs less as we age and is most frequent when traveling, having vivid dreams, or brain injury. Research of déjà vu may prove vital for advancing interdisciplinary research, our interpreting of the world, and answering, arguably, many of life's greatest mysteries: human consciousness and even comprehension of life & death. Societal views of déjà vu have often labeled it abnormal, mysterious, and magical. As déjà vu transitioned from the realms of literature to scientific inquiry in the late 1800s, it captured the interest of the rapidly developing psychological and neurological disciplines. This project seeks to help clarify this issue by exploring how the explanations of déjà vu shifted from literary and artistic interpretations to an empirical research question. In this project, a literature review of historical and contemporary studies of déjà vu will be conducted to help map the conceptual overlaps, theoretical shifts, and methodological progression of research on déjà vu. Understanding how déjà vu's reconfiguration is an empirical question will help us better understand the entangled relationships between the body and experience (i.e., embodied experiences). Additionally, advancing interdisciplinary research about the experience of déjà vu may be vital to advancing phenomenological and scientific (e.g., neuroscience) understanding of many of life's greatest mysteries, from differences in human perception to diverse experiences of lived reality.

Within STEM, physics ranks among the least aligned with the US population regarding racial and gender representation. This not only has the potential to hinder new discoveries and innovations, it also highlights a lack of equitable opportunities for individuals. In an effort to identify ways in which teaching practices may contribute to this problem, our research explores correlations between active learning strategies and growth in students’ conceptual understanding. The data analyzed are from a pre/post survey in a two-year college calculus-based introductory university physics class with a primarily Vietnamese, Black, and white population. We present topics including force and free-fall that show either substantial or limited improvement in student learning gains. We compare data across several demographics, and relate corresponding learning activities. We provide recommendations to improve both learning outcomes and instructional methods, with the aim of increasing opportunities for all identities to complete degrees and pursue career goals.

When students describe physics, they often associate this science with facts, formulae, and objectivity. Unfortunately, all too often, STEM classes completely overlook cultural influence and when it is discussed, it is described as a historical and static phenomenon. This can block students from connecting physics to their own experiences. In our research, we asked students to reflect on their sense of the nature of physics and how their own experiences influence their perception. We used a phenomenological qualitative analysis to investigate 51 students’ ideas across five introductory physics classes at a two year college. Using an emergent theming analysis, we coded students’ written descriptions of physics and how their background shaped their ideas. Students described their own familial, cultural, and professional backgrounds, as well as their instructors’ identities and teaching methods, as impacting their perspective of subjectivity and objectivity in physics. By making space in class to compare and contrast physics culture with students’ own experiences, we hope to show students that their individual background is key to shaping their learning and improving the often inequitable field of physics.

In 2020, the largest protest movement in US history happened in the wake of the murder of George Floyd. In response, the American Institute of Physics (AIP) conducted oral history interviews with prominent African-American physicists, seeking to understand their perspectives on dismantling systemic racism in the physics community. Oral histories provide an alternate view of history that cannot be quantified and exists outside of the normative view of truth; was this an effective strategy for inciting change in a discipline that pedestals objective truths over subjective truths? Moreover, does limiting interviews to notable, outstanding African-American physicists limit the AIP’s ability to capture the range of truths systemically experienced by African-American physicists? This project will investigate the strengths and drawbacks of these oral histories, using primary sources such as National Science Foundation censuses and secondary sources to analyze whether these physicists’ experiences can be generalized and if this generalization holds power over the scientific doctrine of objective physical truths. The treatment of the African-American experience as a monolith dilutes the intentions of the oral histories by withering away intersectional aspects of the experience. As a result, the interviews lack an intersectional and class analysis that reflects how American physics has interwoven with a racial capitalist political economy. By providing an intersectional historical analysis of 20th-century physics, this paper will aim to rethink how large trends are historicized with individual narratives. The framework used to incorporate and critique oral histories in research simultaneously can serve as a model for future history of science research.

I am studying the phonetic effects of childhood exposure to a secondary language on a speaker’s adult primary language. Although some studies explore the effects of secondary language exposure during childhood on the ability to learn that language later in life, there are no studies on the effects of secondary language exposure during childhood on a speaker’s primary language. This study of two English speakers from the Boston area aged 18 and 20 compares the Voice Onset Time of a speaker exposed to Gujarati during childhood to a speaker from a monolingual English household. Voice Onset Time (VOT) is the amount of time between the release burst of a stop phone and the onset of voicing. While American English has a 2 way VOT contrast with no negative VOT (voicing before the release burst), Gujarati has a 4 way VOT contrast with contrastive negative VOT. I recorded the two participants reading aloud English words containing the sounds /p t k b d g/ as the initial consonant.To assess the extent to which sound patterns might differ between participants, I analyzed these sound files in Praat for VOT, for the F1 and F2 at the 25%, 50%, and 75% point of the following vowels, and for the duration of the syllable. The positive hypothesis posits that the speaker with Gujarati exposure would demonstrate significantly more of the negative VOT characteristics typically found in Gujarati speakers than the speaker only exposed to English. These findings may provide insight into both the process of bilingual language acquisition and the long term effects of childhood exposure to secondary languages. In the future, I’d like to expand the study to include more participants and additional speaking or listening tasks.

The crustaceous zooplankton Daphnia magna usually reproduce asexually but are also capable of sexual reproduction. Daphnia populations are generally all female; however, they occasionally produce male offspring for sexual reproduction. In lakes, the production of males and sexual eggs (ephippia) usually occurs in autumn, allowing Daphnia populations to reestablish in the spring. However, in our research, introducing males into some replicates but not others increases variability and decreases the ability to determine treatment effects. There are many hypotheses for what causes female Daphnia to have male offspring – the most common being alterations in light cycles or temperature – but we have encountered male offspring despite controlling these variables. Another hypothesis is that crowding leads to behavioral changes that cause female Daphnia to produce males. I am exploring this hypothesis to determine if the production of males is a behavioral response to crowding or to a lack of food availability. My experiments consist of two treatment groups of four replicates each: crowded and uncrowded. These groups are then fed proportionally upon creation, and reproductive behaviors are measured. The Daphnia cultures are fed with a mixture of three species of algae – Ankistrodesmus sp., Scenedesmus acutus, and Selenastrum capricornutum – to maintain a consistent level of food availability based on measured algal cell density estimated from in vivo fluorescence. If the hypothesis that density causes a behavioral change in reproduction is correct, the crowded treatment will produce more male offspring despite having the same amount of food. If food availability is the major factor impacting reproduction, the number of male offspring between the two groups will be constant. Eliminating sporadic male production would improve reproducibility within and between experiments.

Germany's Energiewende, a pioneering transition to renewable energy, has drawn global attention. Yet, despite initial enthusiasm, disillusionment has grown among energy sector workers and civil society. This study aims to uncover the factors driving this disillusionment. The primary research question is: What contributes to disillusionment amidst the Energiewende transition? Through qualitative analysis of data from the IGBCE union, a key stakeholder in the transition, this research examines the gap between policy objectives and worker concerns. Hypothesizing that a lack of legal support fuels dissatisfaction, the study contextualizes evolving worker attitudes through literature review. Concerns over job insecurity and transition policy inadequacies have shifted sentiment despite initial optimism. Using a comparative case study approach and narrative analysis, I explore how the IGBCE's legal narrative reflects worker grievances. By examining documents from 2015 to 2023, including union materials, recurring themes and narratives are identified. This illuminates the complex relationship between government policies, and worker discontent amidst energy transition. Employing sociological theories such as the "treadmill of production" and "creative destruction," I examine barriers to a just and sustainable transition, questioning whether there are other growth factors that leads to advanced economies being stuck on a "treadmill". Analysis reveals that the Energiewende is primarily driven by societal pressure, prompting political and economic transitions. This shift also influences the stances of labor unions, exemplified by IGBCE's evolution from advocating for fossil fuels to embracing renewables. However, union interests persist in advocating for members. Consequently, with only half of energy jobs governed by collective bargaining agreements aimed at ensuring fair wages, dissatisfaction among workers intensifies. Therefore, workers find themselves vulnerable to precarious conditions and insufficient wages. Result also shows notable lack of government support, further exacerbating worker concerns. This deficiency underscores the importance of unions as effective representatives of workers' interests, with perceived benefits such as collective bargaining power and legal representation, which contribute to the increasing appeal of unions for workers seeking support and protection.

It is well established that formerly incarcerated individuals face stigma upon their return to society, negatively impacting reintegration efforts and life outcomes. Existing literature highlights the significance of stigma in influencing reentry outcomes, emphasizing gender-specific challenges faced by formerly incarcerated individuals, but fails to quantitatively study how these gender stereotypes impact public stigma. Although some research does address variations in the formerly incarcerated population and their influence on public opinion, these studies rarely focus on gender. My study directly explores how gender and stigma interact throughout reentry, by quantitatively investigating public perceptions of formerly incarcerated individuals in Washington State. I develop vignettes about hypothetical formerly incarcerated individuals that vary key facts between participants to examine the extent to which gender, offense type, and post-release behavior influence the public stigma of incarceration. Respondents are randomly assigned these vignettes to evaluate their varying perceptions, unveiling whether gender leads to differential perceptions and therefore, differential treatment in the daily lives of formerly incarcerated individuals. My preliminary expectations are that the public will value relationship-building (a traditionally feminine ideal) more for formerly incarcerated women than for formerly incarcerated men. Additionally, I hypothesize that respondents will exhibit a greater willingness to interact with formerly incarcerated women as compared to their male counterparts, due to negative stereotypes about male aggression and criminality. The implications of this research extend to a broader understanding of the obstacles faced by individuals with criminal records in achieving successful rehabilitation. My analysis will help create more targeted and effective reintegration support policies by uncovering how public stigma impacts the reentry outcomes for women differently than for men. Addressing these dynamics is crucial for fostering inclusivity and dismantling barriers that impede the social and economic participation of formerly incarcerated individuals.

Bilingualism has been shown to have numerous cognitive benefits, ranging from improved executive function to possible protection against neurodegenerative diseases. Structurally, early childhood bilingualism correlates with expanded subcortical regions, specifically the basal ganglia (BG), an area most famously involved in functions such as motor control. In this project, we explore possible domain-general effects of infant and early childhood bilingualism. Specifically, we investigate the effect of early childhood language environments on motor skills in the form of rhythmically controlled babbling. Audio recordings were made in infants’ natural environments using the LENA system. Infants’ own utterances were captured and preprocessed. From these data, I first calculated the ratios of language use in the environment (English versus Spanish), and from these ratios, I categorized infants into monolingual English, monolingual Spanish, and bilingual groups. To determine the rhythmicity of babbling, I isolated subjects’ utterances from the audio data based on criteria that define canonical babbling (e.g., CV syllable structure where C is supraglottal, 4 or more syllables, and so on). For the purposes of this project, we defined the perceptual center as the peak intensity of each syllable and used it to determine the duration between syllables, or inter-syllable duration. The variation of syllable duration was subsequently calculated. We anticipate more rhythmic (i.e. lower variability across inter-syllable duration) babbling in bilingual subjects than in monolingual English and Spanish subjects. However, in accordance with previous findings on early childhood language development, we do not necessarily expect to see differences in whether or not infants are in the canonical babbling stage based on language group. We expect these findings to contribute to the theory of domain-general benefits of early life bilingualism through the quantification of language development milestones.

 The aim of this project is to process and analyze fieldwork data on Triestin to help construct a JIPA (Journal of the International Phonetic Association) Illustration of the language’s main sound systems. Triestin (Glottocode: trie1242) is a dialect of Venetian, a language spoken by approximately 200,000 people in Trieste, Italy. Triestin is spoken at the unique intersection of the Germanic and Slavic language families, and as the number of native speakers of Venetian dialects continues to decrease each year, it becomes increasingly crucial to document the linguistic intricacies of such a vulnerable dialect (UNESCO). Four native Triestin speakers were recorded reading 100 words from a word list. These words elicited vowels in different environments (each vowel sound would be surrounded by other specific sounds), allowing us to record the variations in the pronunciations of each vowel. After evaluating the data from two speakers, Triestin was found to have a 5 vowel system, differing from Venetian and Italian’s 7 vowel systems. Triestin merges the mid-high and mid-low vowels (vowels made with the tongue in the middle of the mouth), a phenomenon not documented in any other Venetian dialect. We found that the two mid vowels vary significantly depending on their environment, raising to [e] and [o] in stressed syllables and lowing to [É›] and [É”] in all other environments. We anticipate finding a system of vowel harmony, whereby the mid-vowels lower in environments to match the height of surrounding vowels. Since Triestin is the only Venetian dialect known to have a 5 vowel system, our goal is to add to the growing body of research on unique vowel systems. In doing so, we investigate the complex relationships between language families and dialects, and provide valuable documentation of Triestin as a low-resource language.

Cleft palate is a prevalant congenital anomaly in the United States, often creating differences in speech and communicative efficacy among affected children. When acquiring speech with a cleft palate, children use compensatory misarticulations to help force air through the mouth instead of the nasal passages. When the cleft is surgically repaired, children continue to use these marticulations. In my undergraduate research, I helped develop a study to assess these misarticulations. Our question was whether the velopharyngeal port (the opening between the oral and nasal passages) was open or closed during these compensatory articulations following palatal repair. I determined a list of words to have children repeat in order to elicit these misarticulations. The data used to answer our research question are novel -- vocal tract MRI (vtMRI). These data show the whole vocal tract moving while a participant is in a standard MRI scanner, reading aloud or repeating phrases. I am thus able to tell for each speaker and for each type of misarticulation, whether the velopharyngeal port is open or closed during misarticulations. This research presents some of the first quantitative data indicating whether misarticulations following palatal repair are habitual or structural in nature, and may indicate pathways for speech therapy to address the misarticulations. 

This study seeks to explore the perspectives of Latine students at the University of Washington (UW) regarding racial representation within the institution. Do Latine students see their experiences and identities represented and reflected in university life? How do Latine students experience and perceive Latine representation (or lack thereof) in terms of the composition of faculty, student body, and community spaces at UW? This research project aims to uncover the realities, challenges, and promise of support and community on campus. Through interviews with Latine students, faculty, and staff, this inquiry will describe how students find support at UW and navigate their academic environment. To attain a comprehensive understanding, the research utilizes a combination of primary and secondary sources, incorporating interviews with Latine students, staff, and faculty at the UW to capture personalized and nuanced perspectives. By examining the lived experiences and perspectives of individuals directly affected, the study aims to thoroughly examine the complex dynamics at play. Exploring the lived experiences of Latine students, the study will contribute to the discourse on racial representation in academia and its impact on student well-being and academic success. The findings will inform discussions on how institutions can foster an inclusive environment that recognizes and supports the diverse backgrounds of all students. This research project has emerged from ongoing conversation and collaboration with the Washington State Commission on Hispanic Affairs members. This research project will inform the community report that the Commission is preparing.

The National Associations of the Deaf astimates there are 308,648 deaf and hard of hearing D/HH students in the US, 79.2% of D/HH students attend mainstream school. D/HH students tend to stuggle academically and socially in high school which leads to low self-esteem. This project uses a Critical Youth Participatory Action Research (YPAR) ethic, framework, and methodology to put power of research and advocary into the hands of the D/HH youth at a PNW high school. YPAR is an epistemological framework in response to the historical dehumanization of marginalized communities. It democratizes ad decoonizes research by privileging the knowledge production of vulnerable communities and foregrounding justive and change. In practice, YPAR develops research skills of typically non-academic co-researchers who drive the research question, design, and analysis. The aim of this project is to guid the students and build their skills to name the challenges they face as minoritized D/HH students, to collect and analyze relevant data, and to use the data to drive policy. the signifiicance of this project is the methodology thatdemands youth voices to be centered in policy making and cultural change. The expected results are findings that speak to the challenges of D/HH students at a PNW high school. The hope is for the D/HH students to present their finingds to policy and decision makers. There is limited research on D/HH students using YPAR methodology and this research will add the perspectives of D/HH students of the discourse.

Vocabulary acquisition in children involves many cognitive processes, with semantic descriptions playing a potentially crucial role in word learning. Broader literature points to a positive impact of metacognitive strategies early in children’s learning. Additionally, studies have highlighted semantic descriptors in facilitating word acquisition. However, there is a lack of literature regarding the direct correlation between children's productions of semantic descriptions and success in word learning, which is the current focus. We examined the number of semantic descriptions produced by children alongside their corresponding word-learning performance to determine whether a relationship exists. We analyzed data from 33 participants ages 2.7 - 6.7 years (mean = 4.6 years, SD = 1.2) with an average Expressive Vocabulary Test-3 standard score of 120. Our team coded a total of 165 word-learning assessments from videos of participants engaging in a storybook listening activity where participants were exposed to novel words paired with objects, assessing their receptive and expressive word learning. Participants were asked to look at the objects, imitate the novel words in real-time, and both point to and label the associated objects after a delay. We recorded the number of times children made semantic descriptions (ex. “The gek is spiky”), total correct identifications (receptive performance), and total phonemes correctly imitated and labeled (expressive performance). For analysis, we tested the correlation between semantic descriptions and both receptive and expressive word learning scores. I hypothesize that a positive correlation between these scores will indicate that children who produce semantic descriptions find more success in word learning. I anticipate that our study will contribute to a better understanding of how semantic descriptions play a role in receptive and expressive word learning outcomes. Also, study findings could serve as a foundation for future research on how parents and educators might better support children’s vocabulary acquisition.

DLD is a highly common yet underdiagnosed neurodevelopmental disorder which impacts receptive and/or expressive language and cannot be attributed to another etiology such as hearing loss or lack of early language exposure. There is a lack of research into the brain differences that characterize DLD, especially in adult populations. The purpose of this project was to explore whether there are brain differences that underlie reading ability in adults with DLD. I specifically focused on the white-matter integrity of reading-related tracts using diffusion magnetic resource imaging (MRI). Participants included a group of adults identified as having DLD, as well as a group of typically developed adults. To measure reading ability, we used the Test of Integrated Language & Literacy Skills (TILLS) and focused on the Nonword Reading and Reading Comprehension subtests. For brain analysis, I first used data-driven hypothesis-free analysis of white matter across the whole brain with tract-based spatial statistics (TBSS). Next, I used probabilistic tractography to evaluate white-matter microstructure in tracts in the dorsal language stream, and in particular, the bilateral SLF. I hypothesized that lower reading scores would be associated with white-matter tract differences in the bilateral SLF. This research matters because reading ability is something that impacts all aspects of modern life, including learning, navigating directions in the world, and succeeding in the workplace. Adults with DLD are often undiagnosed in childhood, and this can carry lifelong social, emotional, and career effects. It is essential to understand whether brain differences underlie DLD and reading, particularly in adulthood, in order to better understand how to ultimately understand, support, and treat this disorder.

Oyster farms sometimes experience large die-offs in the summer, often attributed to warmer temperatures and/or disease. This project aims to investigate to what degree prior stress exposure might influence resilience during subsequent stressors such as those experienced during these large die-offs. For this project, oysters across various life stages including spat (1-5 mm), seed (5-10 mm), large seed (10-40 mm), and adults (40+ mm) were raised at Point Whitney Shellfish Laboratory. Half of the oysters were heated to 25ºC for 6 hours a day for three weeks and mechanically stressed via tumbling for 15 minutes approximately once a week. The remaining, control oysters were held at ambient conditions (17ºC). After seven weeks with both cohorts at ambient conditions, control and stressed oysters were subjected to 30ºC for 30 minutes followed by 15 minutes of tumbling and immediately sampled for RNA extractions. Oysters unexposed to this secondary stress were also sampled. Select genes were characterized to evaluate physiological consequences of environmental hardening and acute stress. Growth rates and survival were characterized for oysters at all life stages. Together the phenotypic and gene expression data will provide insight into environmental memory associated with hardening which has implications for aquaculture and ecosystem function. It is generally expected that exposure to an initial stress will induce a hardening phenotype in subsequent exposure that will be detected using gene expression. Further it is expected that morphology and response phenotypes will vary across life stages. Oysters are the most commonly farmed marine species, though they are threatened by climate change. This study is part of a larger body of research aiming to improve aquaculture and its resilience to climate change. It will help to inform hatchery practices to produce higher yields despite warmer temperatures.

Panãra, an Indigenous language native to Brazil, is currently the focus of Dr. Myriam Lapierre, Sunkulp Ananthanarayan, Ella De Falco, and Jessamine Jeter as some of the only linguists to document and conduct a comprehensive study on this language. Our research focuses on streamlining the process of organizing and analyzing field data – specifically in the context of Panãra, though generally applicable to other Indigenous and/or under-researched languages  – for use in future research by Dr. Lapierre and other scholars in the field of linguistics as it applies to Indigenous and minoritized languages. We have digitized the data from the field journals of Dr. Lapierre and the graduate students working with her, and our current focus is on the analysis of verb and sentence construction, via this digitized data, to organize grammatical paradigms into efficient and accessible indexes. We are also compiling and organizing PDF, image, sound, video, and experimental data for use on the California Language Archive (CLA) with a similar focus on efficiency and accessibility. The completion of this research entails the more complex understanding and organization of Panãra sentence and word structure for use in future research, both by Dr. Lapierre and by other scholars, as well as for usage in a Panãra dictionary. Our expected results also involve the creation and organization of the CLA page dedicated to Panãra, with a transparent structure making this data available to a wider audience of both linguists and non-linguists interested in learning more about the language.

Food insecurity is a global problem with one out of every six people in the United States relying on charitable food assistance organizations to meet their needs in 2022. However, organizations like food banks are underfunded and overwhelmed, relying on donors to continue aid. One food bank in the Seattle area received 70% of their donations from individual donors as financial aid, and a significant contribution comes from grocery stores in the form of food surplus. Previous research has shown that tax deductions motivate grocery stores to donate and limit food waste, however little is known about the care involved in the decision of selecting food to be donated. The quality and quantity of donations varies depending on the store and department. I explore how managerial discretion explains why different grocery store departments choose to donate food or throw it away. I conduct one-on-one structured interviews with three managers in four grocery stores in a Seattle neighborhood to find what factors influence the amount and quality of donations. Additionally, I uncover the internal and external constraints of managers in different departments. Managerial discretion has been studied in various firms yet it has not has not been explored explicitly within the context of donations. Similarly to charitable donations, which are scarce in the context of individuals making philanthropic decisions within organizations. I predict managers’ values and opinions on charity, the grocery store’s philanthropic organizational structure, and the perceived strength of the food bank-to-manager relationship will influence the quantity and quality of donations. I also predict that managers in stores with a higher philanthropic public image will contribute donations of higher quantity and quality. The findings from this study will help identify factors to increase the quantity and quality of donations so that stores can donate higher quality food.

Autism is a developmental disorder that involves differences in social communication skills and the presence of restricted or repetitive interests. One way children learn language is fast mapping: the process of forming associations between new words and their meanings. Growth in language production and comprehension is predictable by 2.5 years of age, and autistic children who learn more slowly during fast mapping tend to score lower in receptive language measures. However, there has not been extensive research on the differences in language learning during fast mapping between autistic and neurotypical children. Therefore, this study addresses fast mapping in one particular learning context and tests whether receptive language learning in a storybook-listening task differs between autistic and neurotypical children. There are 47 participants aged 2.7 to 11 years (mean age = 5.8 years, SD = 2.0), of which 17 are autistic (mean age = 7.8 years, SD = 1.6) and 30 are neurotypical (mean age = 4.7 years, SD = 1.2). I code videos of participants completing the storybook listening task for demonstrations of receptive language skills for novel vocabulary words — namely, the accuracy of participants' responses to prompts to look at named novel objects. I use t-tests to measure the statistical significance of the differences between the groups. Additionally, I conduct group matching based on participants' Growth Scale Value (GSV) scores on the Peabody Picture Vocabulary Test-5 (PPVT), a standardized measure of receptive vocabulary (autistic participants’ mean PPVT GSV score = 488.1, SD = 18.5; neurotypical participants’ mean PPVT GSV score = 479.2, SD = 12.2). I hypothesize that the autistic group of participants has lower receptive language learning scores. I hope the findings of this study can provide insights to help professionals in the field of communication sciences and disorders in their work with autistic children.

This project aims to document the phonology (sound systems) of Triestin, a Venetian dialect spoken by about 200,000 people in Trieste, Italy. Triestin lies at the intersection of the Germanic, Romance, and Slavic language families and thus exhibits many unique phonological phenomena. The documentation of these phenomena is increasingly important as the number of native speakers rapidly decreases. I recorded four native Triestin speakers reading over 300 words each to elicit a variety of words and sentences. In my illustration, I analyze two main phenomena: (i) the vowel system and (ii) intonation. To analyze the vowel system of Triestin, I took 100 words with vowels in different phonological environments (surrounding sounds) and analyzed patterns in their formant values (resonant frequencies that distinguish one vowel from another). I found the vowel system is unique in its reduced size, having only 5 distinct vowels as opposed to other Venetian dialects’ 7 vowels. My analysis describes how this reduced vowel system exhibits variations unique to the dialect, such as raising in stressed penultimate syllables. Triestin sentences also have a rich system of intonation that differs significantly from other Venetian dialects. In my research, I demonstrate how declarative sentences’ intonation is a function of the pragmatics (contextual meaning) of the conversation. The data also suggests that the intonation interacts with the stress of the final word in the sentences, resulting in greater rising intonation for sentences ending in a stressed syllable. The unofficial languages of Italy, including Venetian and its dialects, are extremely under-documented. Projects such as my illustration of Triestin phonology help with expanding the body of literature on the unique features of endangered dialects. This project is also the first step in a more holistic documentation of Triestin, with future projects aimed at studying the syntax and sociolinguistics of the language.

Loudness is a major component of how comfortable someone is with the sounds they hear, and looking at how people perceive the loudness of sounds around them is a crucial part of understanding how people process sound. Previous research by Dr. Yi Shen's laboratory has created an application that allows rapid measurement of loudness across frequencies and levels. The purpose of my project is to expand upon what was found using the previous application. To do so, I am measuring people’s perceptions of loudness across multiple bandwidths. In this project, I guide participants to listen to a sound stimulus in a specific condition, and then select what they perceive the loudness of said sound stimulus to be on an application. Multiple bandwidths of sound stimuli are presented (examples of bandwidths being presented include "Puretone", "1/2 Octave", etc.) and participants then rate how they perceive the loudness of each sound on a scale from "Not Heard" to "Too Loud". By utilizing the application to look at said stimulus bandwidths and performing computer modeling of the results, we gain a greater understanding of how a person’s loudness preferences from one ear can differ across frequency, level, and stimulus bandwidth. We can then analyze these interactions and create a more comprehensive model of the auditory system. I anticipate that results will show participants' bandwidths are consistent with the proposed bandwidths of prior models. This project has future implications regarding rapid identification of issues within a person’s auditory system, how comfortable they are with the loudness of what they hear, and the ability to deliver personalized interventions in assistive hearing devices accordingly.

Religious nationalism is receiving growing attention because of its current influence in democracies, but it manifests itself differently across countries. Previous research has identified socio-historical characteristics that impact the salience of religion as a factor in an individual’s conception of national identity. However, scholars of these studies have mainly used International Social Survey Programme data to analyze European nations. My study uses World Values Survey (WVS) data from 1981 to 2022 to evaluate democracies from a global perspective. I conduct a multilevel analysis to determine the salience of contextual characteristics that prime religion to be used as a vector through which nationalism is mobilized among individuals. Accounting for individual-level factors, I identify which country-level factors influence the relationship between religion and support for nationalism among individuals surveyed. I determine an individual’s support for religious nationalism using the beliefs and groups they express to support in the WVS. I then compare these results to events of national stress to determine if a relationship can be identified. My results suggest that religions favored in church-state relations are more likely to be included in conceptions of national identity. Additionally, affiliates of favored religions are more likely to support nationalism during times of outgroup threat, such as increases in immigration, socio-economic turmoil, and political/cultural shifts. By completing a global analysis of these phenomena, I am able to identify a more comprehensive pattern, something less expansive studies struggle to achieve due to hyper-partisanship debates that can overshadow case studies and regional analyses. Furthermore, by determining which socio-historical characteristics have the greatest impact on conceptions of a religious national identity, I provide a framework to develop a predictive theory on the circumstances under which a religion is not only primed to mobilize nationalist movements, but then comes to be employed to mobilize these movements.

Previous studies have found that jellyfish benefit from warming temperatures and eutrophication, resulting in large jellyfish blooms worldwide. However, the impact of warming ocean temperatures on the cross jellyfish, Mitrocoma cellularia, which are common in the Pacific Northwest, has been relatively unexplored. This study tests the hypothesis that warmer water temperatures would increase M. cellularia growth. A total of 36 individuals were collected from the UW Friday Harbor Laboratories Dock, San Juan Island in September 2023. They were observed in individual jars without water flow, and equally distributed across four separate temperature-controlled baths at 13°C, 16°C, 18°C, and 20°C. Over the 96-hour exposure period, they were fed excess live zooplankton collected before each feeding and had daily water replacements. Surface area was measured with ImageJ, mortality, and overall condition after the acclimation period (initial), day two (mid), and day four (final). A growth rate polynomial regression characterized the final measurements of normal condition jellyfish with a maximum closest to 16°C, resulting in an R2 =0.24 and p-value <0.05. At 20°C, shriveling and mortality increased. While 20°C decreased M. cellularia growth, our results support previous studies that jellyfish are resilient to impacts of climate change within the range of near-future prediction for the Pacific Northwest. Further investigations of the adaptability of M. cellularia to changing temperatures is necessary to understand the future outcomes of their ecology. Examination of this species can inform the state of a jellyfish ecosystem’s adaptability to climate change, which will allow further studies to create solutions to combat changing environments for ocean habitats.

Two-dimensional (2D) materials are layered materials made up of either a single or very few atomic layers. The properties they exhibit are fundamentally different from those of three-dimensional crystals, and they provide us with high levels of control to design novel systems in order to study new physical phenomena. 2D dielectrics are used in electrostatic gating to control the displacement field and carrier doping being applied to a 2D sample, making the properties and performance of 2D dielectrics very important. In 2D heterostructure devices, the standard dielectric used is hexagonal boron nitride (hBN). However, a recently published paper  claims that the novel 2D dielectric Bi₂SeO₅ is readily exfoliated, transferrable, and has a dielectric constant of 15, significantly higher than that of hBN, which ranges between 3-4. In this project I aimed to determine the dielectric constant and breakdown characteristics of Bi₂SeO₅ in hopes of finding a more effective substitute for hBN. I successfully measured the dielectric constant of Bi₂SeO₅ by fabricating a 2D graphene heterostrcuture device and taking graphene transport measurements comparing hBN and Bi₂SeO₅. From the data obtained I found the dielectric constant to be 14.3, which agrees with literary values. I fabricated a Bi₂SeO₅ backgate using a dry transfer technique in order to perform characterize the breakdown characteristics of the dielectric. By applying a voltage to each contact, grounding the gates, and then floating all other contacts, I measured the current passing through the dielectric until we began to see an exponential trend in order to measure breakdown. This is of interest because if Bi₂SeO₅ is proven to be more effective than hBN, it would let us apply a higher displacement field and doping to samples which would let us access new, exotic phases in 2D materials such as WTe₂.

Marine sediments are a critical carbon reservoir, locking both inorganic and organic carbon in sediments for thousands to millions of years. This carbon is released in part through the production of methane at methane seeps, places where methane gas is emitted from the seafloor. In this study, I characterized source differences and quantified the amount of inorganic and organic carbon stored in Puget Sound sediments at a methane seep site. A multicorer collected sediment samples at two sites: one at the Alki Point methane vent field and a control site within the Main Basin of Puget Sound. The sediment cores were subsampled in 2 cm sections downcore and stored for later analysis. Analyses included physical parameters (grain size, percent loss on ignition (% LOI), and 210Pb sediment dating) and geochemical parameters (Total Organic Carbon (TOC), Total Carbon (TC), del 13C TOC, del 13C TC). Both sites were silt-dominated (Alki = 37 microns, Control = 30 microns), with a well-mixed layer 15 cm thick. Organic content (% LOI) was high at both sites (Alki = 7.5%, Control = 6.9%). Likewise, TOC and TC showed trends of increased organic material at the methane seep site. Isotope data indicate that methane-influenced sites have lighter del 13C. Given the amount of carbon they store and the potential for these reservoirs to be disturbed by bottom trawling, deep sea mining, and other invasive human activities, understanding how carbon is cycled through marine sediments is critical for preserving these reservoirs and adequately factoring their role in the carbon cycle into global climate models.

Protists are single-celled eukaryotic plankton that are abundant and diverse in the surface ocean. Many of these protist species are motile often through the use of flagella or cilia. The gene expression of protists is closely synchronized with the daily cycle of light (diel), particularly through the activity of photosynthesis-related genes. However, there has been little investigation of which motility-related genes are expressed in open-ocean protists, among which species, and whether motility-related gene expression is also coupled with the diel cycle. This study aims to investigate the hypotheses that motility-related genes are highly expressed in protist communities, particularly among mixotrophs, and that their expression is synchronized with daytime light. To investigate motility-related gene expression of open-ocean protists throughout the diel cycle, triplicate eukaryotic metatranscriptomes, within the 0.2 – 100 μm size fraction, were collected from 15-m depth approximately every four hours for three days at 158 °W, 41.6 °N, located in the North Pacific Ocean. We annotated protein families (Pfams) present in the metatranscriptomes with Gene Ontology (GO) terms. Pfams were selected based on GO terms related to motility, such as flagella, motility, and cilia. This approach resulted in the examination of the expressions of 21 motility-related Pfams.

Marine ecosystems are currently experiencing changes at unprecedented rates. Implementing camera technologies to visually monitor vulnerable marine ecosystems is becoming increasingly common. Images and video can capture a variety of biological indicators of ecosystem health including species abundances, ecological interactions, and individual health and behavior which can be challenging to assess in other ways. As massive amounts of high-quality photographic data are accumulating, the tools to analyze them are often not readily available to scientists. Therefore, collected images may not be analyzed at the timescales required for scientists to assess and respond to these rapid ecological changes. To address this, we have developed a series of machine learning models that can analyze this surplus of image and video data into meaningful insights for researchers. We built the Subaquatic Ecosystem Analysis and Population Estimation Network (SEAPEN) by meticulously labeling millions of images from oceanographic datasets using a model-assisted labeling protocol. Our original models began with human annotated models, but as accuracy and precision progressed, we were able to rapidly iterate and expand SEAPEN’s capabilities. The processed images are then put through our custom training framework and are run until they output trained TensorFlow and TFLite models. These models are then sent through a series of statistical tests and further quantized until they reach our high set of QA/QC standards. SEAPEN has been tested using a variety of image data from Ocean Observatories Initiative to accurately classify organisms at deep sea ecosystems. SEAPEN is also capable of assessing coral bleaching rates, estimating fisheries stock population size, and quantifying the presence of marine debris. By utilizing large amounts of previously unused public oceanographic data, SEAPEN helps put the tools necessary to process old and new ocean data quickly into scientists’ hands.

Zooplankton are vital to the marine food web, supplying nutrients and energy from primary producers to secondary consumers. During Diel Vertical Migration (DVM), zooplankton travel between depth and the surface during day and night to capitalize on food and avoid predation. This study investigated diel differences in zooplankton community composition at two locations, one exposed and one protected, in the San Juan Channel, WA over four days in September 2023. Zooplankton were collected using net tows from surface waters at both sites during day and night times. Samples were analyzed using a stereoscope and different taxonomic groups were counted. Copepods were the most abundant zooplankton taxa at both locations, with mean abundances up to 1000 individuals per cubic meter. At the exposed site, there was a significantly higher (p<0.05) abundance of zooplankton at night versus during the day. The exposed site had significantly higher diversity than the protected site at night (p<0.05). At both locations, species richness was significantly higher (p<0.05) at night compared to day. The exposed location also had significantly higher richness (p<0.05) compared to the protected location during the day. Our results indicate that zooplankton abundance and diversity in surface waters of the San Juan Channel are controlled by DVM, and differences in locations perhaps due to exposure to different flow regimes. This study reinforces the flexibility of zooplankton community composition and emphasizes the importance of understanding factors that influence changes in the base of the marine food web.

The world’s oceans are witnessing a surge in plastic pollution, a consequence of human activities and the growing urbanization of coastal regions. Urban estuaries are complex habitats that are especially good at trapping sediment, carbon, and pollutants, such as plastics. However, our understanding of the extant of plastic accumulation within estuarine sediments remains limited. I determined the first quantification of the total amount of microplastics (>5 mm) in Puget Sound, WA – a heavily urbanized estuary – and identified deposition hotspots related to current hydrodynamics. To measure plastic concentrations, I collected both shoreline and shipboard sediment samples and density extracted microplastics using an NaI solution. Extracted plastics were counted, sized, and categorized under a microscope. To complement these plastic analyses, energy of the environment was determined using both grain size analysis and extraction of current velocities from LiveOcean, a hydrodynamic model of Puget Sound. I found that plastic concentrations are the highest near land-water interfaces, which are correlated with human population. The dominant source of microplastics came from fibers shed from clothing, giving a well-sorted particle size distribution. Furthermore, using the plastic concentration data I developed a predictive model of plastic distribution that relies on Puget Sound currents and could be adapted for other estuarine systems. Providing a comprehensive analysis of the sources and sinks of microplastics in main basin Puget Sound that can be used to inform preventative management on the negative impacts of urban waste.

Benthic foraminifera are single-celled organisms that are a useful proxy of environmental conditions. They can provide insights into water quality, sediment composition, and ecological changes. In the past, urban bays of Puget Sound developed chemical toxicity in sediments surpassing acceptable state limits. Recent mitigation efforts have brought most chemicals within limits. However, foraminifera health parameters continue to show signs of poor environmental conditions including increased test dissolution and decreased species richness and density. Studies on individual embayments are necessary to understand local variations and utilize foraminifera in an effective monitoring program. In this study, we investigated how the percentage of dissolved foraminifera varies based on distance from shore in Elliot Bay, Seattle. We obtained grab samples through the Washington Department of Ecology collected in 2021 from multiple sites in Elliot Bay. We processed the samples and picked up to 300 foraminifera from each sample then calculated the percentage of dissolved Elphedilla hannai, the most common calcareous species in Elliot Bay. Additionally, we reviewed foraminiferal densities, species richness, and diversity for distance from shore. We found that the percentage of dissolved Elphedilla hannai increased closer to shore and with decreased depth. These results suggest a spatial influence on the health of foraminifera.  As Elliot Bay is a major maritime hub, its shorelines are heavily industrialized, and areas both shallower and closer to shore may be experiencing higher levels of anthropogenic activities. This study highlights potential water quality gradients in urbanized embayments such as Elliot Bay, contributing to our overarching aim of understanding interactions between foraminiferal health and environmental factors in urbanized embayments.

The invasion of Indo-Pacific lionfish (Pterois volitans) throughout the western Atlantic, the Gulf of Mexico, and the Caribbean has led to declines in the biodiversity, biomass, and abundance of native coral reef fishes. These ecological consequences have been thoroughly documented in shallow zones of the reef; however, little is currently known about how lionfish interact with deep-reef ecosystems, and which native fish are the most vulnerable to predation. Deep-reefs may serve as refuges for lionfish, as they would avoid being speared by SCUBA divers in the top 30 meters. Thus, deep lionfish populations could potentially serve as a near-infinite source of replenishment for shallow populations. The microchemistry of calcium carbonate fish otoliths has emerged as a powerful tool in examining fish ecology, including their vertical movement. The elemental compositions of these “ear stones” reflect the chemistry of the water in the fish’s environment, ultimately providing insight into natal origin and dispersal events. I am analyzing the oxygen isotopic signatures in a sample of 10 lionfish otoliths collected across the entire depth range of reefs in Curaçao using micromilling and isotope ratio mass spectrometry (IRMS). Specifically, variations in δ18O from the core to the rim of the otolith indicate changes in water temperature, and presumably depth, from settlement to adulthood. This approach provides insight into whether lionfish migrate between deep and shallow reefs throughout their lives or remain in their deep refuge, how frequent and long these migrations are, and ultimately whether they are susceptible to surface removal by spearfishing. Effective management plans will require a holistic understanding of this invasive predator’s vertical migration patterns and where they are on the reef at various life history stages, and a deepened understanding of the connectivity between deep and shallow populations can inform future research to control mesophotic populations.

Climate change is increasing glacial melt worldwide, causing freshening events in marine ecosystems that rapidly decrease salinity. In the Salish Sea, summer low salinity events occur regularly from the Fraser River meltwater. This negatively impacts many organisms including sea urchins, which are resilient to other climate change impacts like marine heatwaves and reduced food availability. Previous studies found that low salinity impacts tube foot mobility and function in green sea urchins (Strongylocentrotus droebachiensis). We replicated the Salish Sea freshening events with a treatment of 20 PSU saltwater (control 30 PSU) to investigate the impacts on red sea urchin (Mesocentrotus fransiscanus) coordination and spine movement. This study implemented new techniques to quantify urchin motility to determine the negative effects that low salinity had on sea urchins over 96 hours. We quantified spine movement using image analysis to track individual spines of urchins in low and regular salinities. We also measured righting time, which is an indicator of urchin coordination. We found that urchins exposed to low salinity had significantly longer righting times and less spine movement overall. The low salinity treatment spine movement and righting time were highly correlated (p < 0.01), but not in the ambient salinity tanks (p > 0.01). We found that lower salinity waters have the potential to reduce urchin coordination and movement, which may impact urchin populations and kelp forests in the changing climate. Thus, there may be a lessened impact on the kelp forests needed to maintain ocean health.

Foraminifera, a diverse and widespread order of shelled marine protists, can be utilized as a reliable measure of marine ecosystem health due to their innate sensitivity to environmental changes. This study takes advantage of foraminifera to investigate the health of Commencement Bay, which is located in the South Puget Sound and encompasses the city of Tacoma. It is a highly industrialized embayment that features many man made inlets. Commencement Bay now hosts numerous proposed and implemented clean-up efforts, our primary focus being the U. S. Environmental Protection Agency Superfund sites put in place in 1992. This study, part of the Puget Sound Foraminifera Project at the Burke Museum, aims to build upon previous efforts to monitor the health of Commencement Bay and determine the effect of these clean up efforts. Sediments were collected by the Washington Department of Ecology from multiple sites in Commencement Bay and brought to the Burke Museum, where they have been washed, dried, floated, and picked to collect the foraminifera. Where possible, a minimum of 300 individuals was picked for analysis of assemblages. Results varied by location, with low diversity assemblages near the shore and higher diversity assemblages deeper into the bay. This indicates despite restoration efforts, waterways close to shore are still impacted by anthropogenic influences despite mitigation efforts.

Kelp forests are hotspots for productivity and biodiversity in marine ecosystems. Currently along the West Coast, many kelp forests face significant threats due to climate change. Several studies have looked at the effects of ocean warming on kelp, but less is known about the effects of ocean acidification (OA), and how OA and warming in conjunction impact kelp. With climate change continuing to worsen, it is crucial to understand how these drivers may contribute to kelp’s decline. We examined the effects of warming and acidification, both separately and concurrently, on bull kelp (Nereocystis luetkeana) from two populations in Puget Sound, WA. Bull kelp, the primary canopy-forming kelp species in Puget Sound, has a heteromorphic life history with macroscopic (sporophyte) and microscopic (gametophyte) stages. Little is known regarding how the microscopic stages will be impacted by multiple stressors in Puget Sound. This study aims to better characterize the effects of warming and OA on Puget Sound bull kelp throughout its microscopic life stages. We exposed kelp gametophytes from two genetically distinct populations to elevated levels of temperature and pCO2 in laboratory conditions for five weeks, monitoring their survival, development, and reproductive success from spore settlement to the juvenile sporophyte stage. Our collection sites differed in the overall temperature, urbanization level, and state of the kelp forests. Previous studies have shown declines in reproductive metrics when subjected to thermal stress, and an increased thermal tolerance when subjected to acidic conditions, which is what we expect to find within our experiment. Understanding local adaptability is critical in predicting future population responses to climate change. The potential range shifts or local extinctions that could occur because of these stressors will need to be understood to plan for conservation, restoration, and recovery of healthy kelp forests.

Acorn barnacles (Balanus glandula) are among the most common species found in the Northern Pacific Ocean, inhabiting rocky surfaces across intertidal zones. These barnacles’ distribution is attributed to various factors, including surface exposure, spatial competition, and predation. The size and abundance of barnacles may serve as an indicator of the primary abiotic and biotic processes that influence barnacle populations in highly dynamic intertidal systems. This study tests our hypothesis that higher abundance and smaller-sized barnacles will be observed at higher tide elevations, while lower abundance and larger-sized barnacles will be observed at lower intertidal elevations on the shore. In September 2023, at Friday Harbor, (San Juan Island, WA) we photographed 10 quadrats (24x24cm) at low (0m), mid (1m), and high (2m) elevations. We observed barnacle distributions at two sites along the San Juan Channel. Barnacle abundance and size were measured with ImageJ. We calculated the mean and median barnacle size at each elevation and location. Small barnacles (0-1.5mm) were predominant in the high elevations, whereas large barnacles (>3mm) dominated the lower elevations. Our results are associated with a combination of abiotic and biotic factors. Abiotic factors include greater desiccation risks in the high intertidal elevation. Biotic factors involve food availability, with larger barnacles having greater access to the nutrients coming from the ocean, along with spatial competition, where higher abundance may lead to increased competition, thereby reducing available resources for each individual. Differences between sites may be attributed to diverse characteristics unique to each location.

Growing environmental stresses such as ocean warming have led to a rise in coral bleaching events. Where reef-building corals lose their symbiotic algae that usually supply most of their energy. Bleaching events have led to widespread coral starvation and death, damaging the structure of coral reefs. This study aims to identify if the sex of a coral colony will impact the colony’s ability to withstand growing environmental stresses. Although most coral species are hermaphroditic, we worked to gain insight into the responses of gonochoric corals (in which individuals are either male or female). We focused on a major reef-building species, Porites compressa, from Hawai'i, to better understand how reproductive development compares to bleaching events, and colonies’ ability to recover from bleaching. We hypothesized that female colonies would be better equipped to withstand environmental stress due to excess nutrients stored in their eggs, which can be re-absorbed during stress. While males grow faster at the cost of investing less nutrients in sperm leaving them vulnerable to heating events. Coral polyps from male and female colonies were collected at different points between 2021 and 2023. Using the polyps, I determined each colony’s sex and the developmental stage of each egg or spermary. With this data the varying developmental stages were compared, to present the expected differences between male and female gamete development. I also measured each colony’s annual growth through skeletal growth rings preserved in frozen colony fragments. We analyzed this data to determine if females grow more slowly to better survive bleaching due to earlier annual investment in nutrient-rich eggs, to keep available during bleaching events. Through this study, a better understanding of coral development and success based on the reef’s sex will allow greater predictions to be made in future research as ocean warming increases.
 

Marine debris, classified as solid, man-made litter and material that has been lost or discarded in the ocean, is a persistent pollution issue in coastal regions around the world, and Puget Sound is not an exception. This research aims to investigate the distribution and abundance of marine debris throughout various regions of Puget Sound over time. Since 1987, the Washington Department of Fish and Wildlife (WDFW) has conducted annual trawls to assess bottom fish populations in Puget Sound. The contents of these trawls provide valuable representation of the soft-bottom habitat, including the organisms and debris that inhabit the seafloor. The WDFW’s extensive records of these surveys include the region, depth, type, and abundance of debris collected in each trawl. Using this data, I am exploring patterns in how different types of debris are spatially distributed, examining trends in abundance over the last two decades, and identifying hotspots for debris accumulation in Puget Sound. Through the use of R programming, I conduct analyses and fit statistical models to better communicate my findings. The results of this study provide a better understanding of the dispersal of aluminum, plastic, glass, fishing gear, and other debris that lies at the bottom of Puget Sound. Awareness of these patterns is vital to informing effective clean up and guiding prevention efforts to make community waters cleaner and safer for both humans and marine life.

Antarctic blue whales (Balaenoptera musculus intermedia) are the largest animal on Earth, but much of their life history remains a mystery, including mating behavior, gestation periods, and early development. Antarctic blue whales were hunted excessively throughout the 20th century and are currently listed as endangered due to this exploitation. More than 340,000 blue whales were killed which led to an immense amount of data regarding the length, sex, and reproductive status of individuals. Using this data, compiled by the International Whaling Commission, I will build a model of Antarctic blue whale fetal growth and calf development. I will fit this model to the fetal and calf catch data to predict fetal and calf lengths in each month of the year. This model will result in predictions of the time of year that Antarctic blue whale calves are born and weaned as well as their growth rates. Determining the trends of fetal and calf growth in Antarctic blue whales contributes to the knowledge of reproductive behavior and growth trajectories. This is especially important for Antarctic blue whales, where no breeding areas have been identified. Understanding when calves are born and weaned can be combined with other data, such as satellite tracking, to provide insight into breeding areas.

The Climate-Ready Landscape Plants Project (CRLP) evaluates landscape plant performance under different irrigation treatments across six different geographic and climatic regions. Under the CRLP, four overlapping taxa Physocarpus ‘Diabolo’, Physocarpus ‘Little Devil’, Cercis occidentalis, and Cercis canadensis were chosen to investigate whether xylem anatomy changes in response to water deficits across three different planting sites: University of Washington, Oregon State University, and Utah State University. At each site, 24 replicates per taxa and 8 plants per irrigation treatment were tested under a two-year trial period. In year one, plants were established using the same water treatment. During year two, plants were introduced to irrigation treatments of 20%, 50%, and 80% based on the Water Use Classification of Landscape Species (WUCOLS). Plants were randomly assigned a position in the plot 2m away from each other under one of the three irrigation treatments. Stem cross sections were stained and imaged with vessel sizes analyzed using ImageJ. I expect both Cercis canadensis and Cercis occidentalis to display smaller vessels per xylem area reducing vulnerability to drought induced cavitation across the three treatments. I expect Cercis canadensis to display stronger acclimation to drought conditions because of its high tolerance for cold temperatures across the three sites. I expect Physocarpus ‘Diabolo’ and Physocarpus ‘Little Devil’ vessels to display lesser plasticity in xylem size and density across the three treatments and sites. Compared to the Cercis genus, Physocarpus is ornamentally selected and hybrids of the Physocarpus genus experience reduced water use efficiency with a lesser ability to acclimate to drought conditions. Most plants used for horticulture have not been tested under different irrigation treatments. This research can promote the production and utilization of low water-use plants to create well adapted landscapes under a changing climate for horticulture efforts on a local and industrial scale.

Leigh Syndrome (LS) is a neurodegenerative disease due to the dysfunction of mitochondria. It usually begins in infancy and its incidence is around 1 in 40,000 individuals. Children with LS experience a progressive decline in their cognitive and motor functions often accompanied by severe treatment-resistant epileptic seizures. Mutations in Ndufs4, the gene that encodes a subunit of mitochondrial complex have been linked to LS. Mice carrying Ndufs4 recapitulate several key characteristic clinical manifestations of LS. Using these mouse models, our lab has demonstrated that GABAergic interneurons play an important role in the pathophysiology of LS. Mice with Ndufs4 knockouts (KO) restricted to GABAergic neurons located across all brain regions exhibit seizures. However, seizures in epilepsy patients and animal models typically originate from forebrain structures. Therefore, in this project, we examined whether the inactivation of Ndufs4 in GABAergic neurons of the forebrain alone is sufficient to cause seizures in mice. Homozygotes floxed Ndfus4 mice were crossed with Dlx56Cre+ or Gly2TCre+ mice to KO the gene specifically in interneurons of the forebrain or brainstem. We hypothesized that only mice with KO in the former region will exhibit seizures. Conditional KO mice from these two lines were tested for thermal seizure susceptibility. Surprisingly, both Dlx56creKO and Gly2TCre KO mice exhibited thermally induced myoclonic and generalized tonic clinic seizures. These findings indicate that GABAergic interneurons regions outside of the forebrain are critically involved in the pathogenesis of epilepsy in LS.

Rare Intracranial Tumors (RIT) are a heterogeneous group with unmet medical needs. Although infrequent in individuals, RIT affect millions of people who lack effective disease monitoring and treatment. Frequent chromosome gains and losses are common in cancer, leading to the upregulation of oncogenes and downregulation of tumor-suppressor genes, respectively. Somatic Copy Number Variations (CNVs) affect a greater fraction of the genome than single nucleotide polymorphisms (SNPs) and have been correlated to drug resistance and tumor progression, highlighting a potential prognostic value. Cell-free circulating tumor DNA (ctDNA), which are DNA fragments released by necrotic or apoptotic tumor cells can act as a noninvasive cancer biomarker, offering a potential alternative to invasive tissue biopsies. In the present work, we aim to establish the somatic CNV signature of tumor and matched ctDNA to identify non-invasive tumor-related CNVs that may serve as biomarkers for use in liquid biopsy. We performed Whole Exome Sequencing (WES) in gDNA isolated from tumor tissue and matching ctDNA of 15 patients with RITs (pituitary tumor (n=9), craniopharyngioma (n=2), and meningioma (n=4). Raw reads were assessed for quality (Trimmomatic, FastQC), following alignment against human reference genome GRCh38 (BWA). Aligned reads were sorted and subject to duplicate removal using Picard. CNV profiles will be generated using CNVkit tool. Data analysis and visualization will be performed using R and python. The most promising aspects of liquid biopsy in cancer applications are cancer screening and early diagnosis because they can lead to better survival results and less disease burden. At the end of this work, we hope to identify the CNV signatures shared between tumor tissue and ctDNA, provide novel insights into the pathophysiology of these RITs and ultimately, suggest promising biomarker candidates for liquid biopsy.

Parkinson's Disease (PD) is a progressive neurodegenerative disease characterized by slowness or stiffness of movement and cognitive impairment. PD is characterized neuropathologically by Lewy Body (LB) aggregates that include lipids, proteins and oligomerized alpha-synuclein. Mutations in the gene glucosidase, beta acid 1 (GBA), are not only the most common genetic risk factor for PD but also accelerate the progression of the disease. We hypothesize that mutations in GBA may mediate faster spread of pathogenic protein aggregation from neuron to neuron. Our previous work has implicated GBA in extracellular vesicle (EV) regulation, suggesting a non-cell autonomous mechanism for GBA accelerating propagation of LBs. To test this hypothesis, we are first exploring how GBA influences EV biogenesis in neurons and astrocytes by examining endolysosomal trafficking in GBA mutated neurons and astrocytes, as well as controls, differentiated from human induced pluripotent stem cells (iPSCs). Our initial results indicate that neurons heterozygous for a GBA null mutation have impaired endolysosomal trafficking with enlarged early endosomes and lysosomes, while astrocytes heterozygous for GBA null do not have impaired early trafficking. These results suggest that GBA mutations differently affect different cell types in the brain and improve our understanding of how GBA influences the spread of LB pathology. I image the iPSC derived neurons and astrocytes using a confocal microscope, for endolysosomal markers and distributions. The goal of this work is to identify novel therapeutic targets for slowing PD progression. 

Parkinson’s disease (PD), the second most common neurodegenerative disorder, is characterized by Lewy bodies, pathogenic protein aggregates that include alpha-synuclein oligomers. The missense mutation p.G192R in the RAB39B gene was recently found to cause X-linked dominant PD. Loss of function mutations in RAB39B are associated with X-linked intellectual disability and autism spectrum disorder. RAB39B is a member of the human Rab GTPase family which plays a role in early autophagosome formation and is implicated in intracellular vesicular trafficking. This project investigates how defects in endolysosomal trafficking caused by the p.G192R mutation in RAB39B gene leads to parkinsonism and neurodegeneration. Because RAB39B is highly conserved, we developed a Drosophila model as human RAB39B and Drosophila RAB39 share 75% similarity in amino acid sequence, including 100% identity at p.G192 and flanking amino acids. Using CRISPR/Cas9 genome editing, we created a RAB39G196R Drosophila model that we are currently characterizing for possible neurodegenerative phenotypes. We are examining locomotor deficits and lifespan in RAB39G196R mutant flies compared to isogenic controls, as well as protein aggregation by Western blot. Complementary to the Drosophila model, we developed a human neuronal model by generating induced pluripotent stem cells (iPSCs) from peripheral blood mononuclear cells (PBMC) of an affected male and similar age unaffected male family member kindred with X-linked PD due to the p.G192R mutation. We are investigating endolysosomal trafficking defects in neurons differentiated from iPSCs using antibodies specific for early and late endosomes and lysosomes. We are also examining whether insoluble ubiquitinated protein aggregates and oligomerizes alpha-synuclein are present in RAB39BG192R neurons compared to control neurons. Understanding mechanisms underlying the pathogenesis of X-linked Parkinson’s disease will elucidate the development of PD and potential novel therapeutic targets.

An important part of addiction recovery is degrading high value associations between drug cues and the drugs themselves. Dopamine plays a crucial role in learning, and is specifically implicated in the prefrontal cortex (PFC) and reversal learning - learning to update and change behavior when it is no longer being rewarded. Past studies have reported elevations in dopamine during contingency reversal, but the timescale of how activity of PFC dopamine neurons maps to reversal learning remains unclear. Here we investigated the activity of PFC dopamine during reversal learning in a longitudinal fiber photometry study, recording dopamine signal on a timescale of seconds. Mice were trained on a reversal learning task where they initially learned that two of four presented odors precipitated a sucrose reward in 85% of the trials while the remaining two odors precipitated the reward for only 15% of the trials. Once the learning was stable, reward probability flipped for two odors (one 85% odor and one 15% odor) and the mice had to update their behavior to the new odor/reward structure. Fiber photometry recordings were conducted during pre-reversal, reversal, and post-reversal stages of the study. Our data replicate findings demonstrating elevated dopamine release during the reversal period, centered around the 15-85 cue. Analysis of the relationship between the dopamine signal and behavior also revealed significant cue, reward prediction error, and 15-85 reversal coding in the majority of animals, suggesting a multi-faceted role for dopamine in the PFC. Given this, dopamine in the PFC may play an important mediating role in the enhancement of associations between drugs and drug cues, but does not play a clear role in contingency degradation.

Neurons, the fundamental cellular units of the human brain and nervous system, are crucial to transmitting signals throughout the human body. In human anatomy, anything that obstructs the communication between neurons could lead to neurodegeneration, decline in function, disorders, and diseases. Alzheimer’s disease (AD), which affects more than 6 million people across the nation as of 2023, is one such disorder. An extensive array of research has been done investigating the underlying cause of the neurodegeneration that occurs in AD. One such theory of neuronal dysfunction, the amyloid hypothesis, points to an accumulation of a protein called beta-amyloid that is present in the brain but in some people accumulates in excess and disrupts neuronal signaling. This ultimately leads to neurodegeneration and cognitive decline. I take part in conducting the AHEAD study, a Phase 2 clinical trial, which is currently underway investigating this theory and a new drug called lecanemab. The drug is designed to remove beta-amyloid from the brain and prevent further neurodegeneration in individuals who have accumulations of amyloid and are at risk of developing AD. This is one of the first preventive clinical trials for Alzheimer's. In this review, I have explored findings from Phase 1 of the AHEAD study and described the screening process for participants for the Phase 2 trial. I have also explored the science behind beta-amyloid, Alzheimer’s disease, and treatment with lecanemab. I expected a large population to pass the screening process, but the statistics show otherwise. I dive into why this is in this project.

The expression of prosocial behavior holds significance across various species, and its disruption stands out as a characteristic symptom in numerous neurodevelopmental and neuropsychiatric disorders such as autism, schizophrenia, and depression. Neuronal activity in the nucleus accumbens, particularly modulated by the neurotransmitter- dopamine, plays a pivotal role in regulating reward-motivated behavior towards both food and social interaction. Although the influence of dopamine on social behavior is extensively researched, there exists a notable gap in our understanding of neuronal activity within the nucleus accumbens during social interactions. In this study, a viral vector was employed to induce the expression of a genetically encoded fluorescent calcium sensor, coupled with mini-scope imaging, to observe calcium dynamics in the nucleus accumbens. The recording of calcium dynamics occurred during an operant task where an experimental mouse pressed a lever to interact with a social partner or in separate sessions to receive a food reward. Our investigation revealed distinct subpopulations of neurons within the striatum that specifically encode food reward, as opposed to other discrete cues or actions associated with the task. The insights gained from this research contribute to a more comprehensive understanding of how signals related to general reward-motivated behavior and prosocial behavior may contribute to disruptions observed in social behavior, particularly in disorders characterized by impaired prosocial behavior.

Dravet Syndrome (DS) is a severe form of childhood onset epilepsy occurring in about 1 out of 16,000 births. The disease is characterized by treatment-resistant seizures, ataxia (or loss of muscle coordination), developmental delay, cognitive impairment, and increased rate of premature mortality mostly due to sudden unexpected death in epilepsy (SUDEP). DS is predominantly caused by a heterozygous loss-of-function mutation in the SCN1A gene, which codes for the pore-forming alpha subunit of the NaV1.1 voltage-gated sodium channel. Our lab has previously shown that selectively introducing these mutations into neurons expressing the neurotransmitter GABA, specifically GABAergic interneurons of the forebrain, is sufficient to cause DS phenotypes in mice. In this study, we investigated whether an SCN1A gene replacement therapy precisely targeted to this interneuronal population can rescue epilepsy and SUDEP. Our lab, in collaboration with the Allen Institute, has developed a novel dual SCN1A-intein-AAV with forebrain GABAergic interneuron targeting capability using a Dlx56-based enhancer. Mice treated with this vector at postnatal day (P) 0 via intracerebroventricular injection were monitored for spontaneous mortality up to P70 and tested for susceptibility to thermally induced seizures. All untreated mice (n=31/31) died by postnatal week 6. In addition, 86.5% (n=13/15) of them exhibited thermally induced myoclonic seizures (MCS) and 100% (n=15/15) of them showed generalized tonic clonic seizures (GTCS). In striking contrast, none of the treated mice died (n=9/9, p=3.3e-14, Fisher’s exact test) nor exhibited MCS (n=0/9, p=7.1e-4, Fisher’s exact test) or GTCS (n=0/9, p=1.1e-5, Fisher’s exact test). These findings suggest that precision therapy targeting the very site of disease etiology can completely protect against epilepsy and related mortality in DS.

The use of opioid drugs for pain management in postoperative settings has been widespread since the 1860s. However, the risk factor for developing an opioid use disorder (OUD) has increased substantially with its continued use, with addiction rates of more than 10% for those taking opioids. An obstacle to abstinence in opioid addiction are the adverse side effects that occur during cessation of drug use once dependence has formed, including nausea, anxiety, vomiting, and depression. The Nucleus Accumbens (NAc) is part of the mesocorticolimbic reward pathway. Decades of pharmacological studies demonstrate that nearly all abused drugs evoke dopamine release within the NAc, thus altering innate systems for how reward is processed. The activity of NAc neurons is strongly regulated by efferent excitatory input from numerous brain regions. The paraventricular thalamus (PVT) a relatively understudied brain region, regulates behavioral responses to reward and aversive stimuli as well as to drugs of abuse such as morphine. Our preliminary data demonstrate that the activity of these projections is highly regulated by the Cannabinoid 1 Receptor (CB1), which mediates the primary psychoactive effect of cannabis. This is particularly relevant on account of recent clinical findings demonstrating that activation of CB1 can ameliorate the aversive effects of opiate withdrawal. Using fiber photometry (which uses fluorescence emission of the calcium sensitive fluorophore, GCaMP, as a proxy measurement for neural activity), I have shown that this circuit is activated by aversive stimuli and inhibited by rewarding stimuli. Furthermore, treatment with morphine can attenuate the pain-induced activation of this circuit. However, whether cannabinoids can influence this circuit's activity to reduce withdrawal symptoms remains untested. Our research will contribute to our understanding of the neurophysiological basis for opiate withdrawal and how cannabinoids could represent a novel class of therapeutics for the treatment of opiate use disorder.

Addiction is characterized by the compulsive use of substances despite adverse consequences, a process closely linked to dopamine-induced changes in the Nucleus Accumbens (NAc) and its role as the brain's "reward center." The NAc integrates information from various brain regions, including the Paraventricular Thalamus (PVT), to produce motivated behaviors. Recent studies have identified the PVT, especially its anterior segment (aPVT), as a critical hub in addiction neurocircuitry, but findings have been inconsistent, likely due to the PVT's heterogeneity and the specific neurochemical and anatomical properties of its connections to the NAc. Prior research has shown that aPVT neurons, identifiable by neurotensin expression, send excitatory projections to the NAc, which are modulated by endogenous cannabinoids (eCBs). These interactions suggest a complex regulatory mechanism. Preliminary experiments used techniques including transsynaptic viral tracing and in vivo calcium imaging, to study the activity dynamics of NAc neurons, particularly those expressing Proenkephalin (PENK) and receiving aPVT inputs, during reward-seeking tasks. I propose to extend these findings by employing a multidisciplinary approach that combines experimental neuroscience with sophisticated computational analysis. By applying dimensionality reduction techniques, clustering algorithms, and machine learning models to neural and behavioral data, I aim to map the functional connectivity within the NAc and elucidate the roles of specific neuronal ensembles in reward-seeking behavior. This comprehensive analysis will not only clarify the neurobiological underpinnings of addiction but also contribute to the development of targeted therapies for addiction and related disorders, leveraging the unique intersection of computational neuroscience and behavioral analysis.

Light sheet imaging differs from other forms of microscopy by illuminating the entire plane of an ex vivo extracted brain sample and computationally compiling these planes post-hoc into a 3-dimensional image. This technique allows users to preserve the structural integrity and map biological units such as cells, axons, and genes onto a standardized brain atlas. Using the SHIELD clearing, preservation, and staining protocol, we prepared fluorescently labelled ex vivo whole, intact brains for light sheet imaging and used the ClearMap open source pipeline on the Hyak cluster operated by University of Washington-IT to overcome computational limitations. While ClearMap registers images and identifies cell counts per brain region, it lacks quality assurance metrics as well as figures for visual confirmation of preprocessing performance. To overcome these deficits, we have built a quality assurance package for ClearMap that includes registration validation and group-level quantification figures. In testing this package we analyzed multiple mouse brain samples exhibiting Fos expression patterns under varied conditions. Using this dataset we found that registration performance between expected and actual slices measured in F1 score averaged 0.7 in medial sections of the brain and we were able to visually inspect the alignment of subregions identified in a standardized brain atlas. Additionally, we created an interactive script to visualize heatmaps that indicate expression patterns for user-defined brain regions on an animal level. This work will enable rapid validation of collected data and third-party package performance while streamlining the analysis, interpretation, and visualization of results. In the future we intend to include useful statistics (one-way ANOVA, classification models, etc.) to provide additional functionality and further increase the speed and quality of data analysis.

In recent years, the United States has seen an increasing number of opioid overdoses, causing many research studies to focus on opioid drugs of abuse. However, we still lack a fundamental understanding of the opioid receptor that these drugs bind to. The mu-opioid receptor (MOR) is involved in both the pain-relieving and euphoric effects of opioid drugs. The Ventral Tegmental Area (VTA), known to be a major source of dopamine, contains a significant amount of inhibitory gamma-aminobutyric acid (GABA) neurons that express MORs. Previous research has shown that activity at the MOR in the VTA is rewarding and that dopamine projections from the VTA to the Nucleus Accumbens (NAc) are activated to reinforce behavior. However, whether the expression of MORs on GABA neurons in the VTA is important for influencing VTA to NAc dopamine activity during reward seeking behavior is unclear. To investigate the role of the MORs in reward seeking behaviors, I knocked out MORs in the VTA and used dLight, a fluorescent dopamine sensor, to measure dopaminergic release from synapses in the NAc during Operant and Pavlovian conditioning tasks in the absence of these receptors. Pavlovian conditioning utilizes chambers in which a house light turns on and a sucrose pellet is delivered via food hopper to food-restricted mice. In the operant conditioning task, mice must learn to complete a nose poke in order to receive the sucrose pellet reward. These tasks allow me to gain a more complete understanding of how the absence of MORs in the VTA changes behavior and dopamine activity during reward-seeking. This research furthers our understanding of how the MOR affects natural reward and motivated behaviors and is crucial in helping us understand how opioids of abuse alter existing brain circuitry to cause opioid use disorder.

 The locus coeruleus (LC), a nucleus in the pons of the brainstem, is the primary source of norepinephrine (NE) in the brain and is responsible for regulating arousal and stress responses. The LC also has roles in pain, avoidance and reinforcement learning, exploration, and feeding. How LC neurons are mediated to generate these distinct responses has yet to be defined. One area of interest is the peri-LC, which sends local GABAergic projections to the LC and is thought to modulate physiological arousal through direct inhibition of the LC. The peri-LC is composed of various populations of GABAergic neurons that may contribute to the inhibition of the LC. We were interested in exploring the role of neurokinin1/substance P, nociception, and enkephalin GABAergic neurons. Using vGAT-cre, Tac1-cre, Noci-cre, and Penk-cre mice strains, we injected an excitatory DREADD into the peri-LC to be able to activate genetically defined neural subpopulations within the peri-LC. I ran the mice through open field and elevated zero maze testing to assess anxiety levels. The activation of all peri-LC^GABA neurons led to a decrease in anxiety-like behavior, which is consistent with an inhibition of the LC and a decrease in LC^NE release. In contrast, the activation of peri-LC^Tac1 neurons increased anxiety-like behavior, while the activation of peri-LC^Noci and peri-LC^Penk neurons had no behavioral effect in these assays. Together, these results suggest that the different peri-LC^GABA subpopulations likely represent distinct nodes that differentially integrate novel stimuli to influence distinct behaviors. Understanding how the peri-LC regulates LC function can give insight into the neurobiological basis of arousal and stress-related disorders, which informs our understanding of neuropsychiatric pathologies.

Reward-seeking behavior, crucial for survival, is hijacked by various neuropsychiatric disorders, notably substance use disorder. Prior work implicated reward-seeking with the dorsomedial striatum (DMS), a brain structure expressing the endogenous opioid dynorphin (dyn) in 50% of its neurons. Previous studies also linked dyn, signaling through the kappa opioid receptor (KOR), with escalation and reinstatement in drug-seeking, however, the exact mechanisms of dyn-KOR signaling remain unknown. Recent experiments from the Bruchas lab show that supraphysiological release of dyn in the DMS with excitatory optogenetics elicited reward-seeking behavior, demonstrating that dyn-KOR is sufficient for reward-seeking behavior, but not that it is necessary. To provide evidence of necessity, I plan to inhibit dyn-KOR signaling in the DMS by expressing the novel inhibitory optogenetic tool Platynereis dumerilii (PdCO) that we recently showed to be useful for such studies (Wietek et al., Nat Methods, 2024). PdCO is expressed in a target neuronal population and begins inhibiting under 473 nm light. Hence, in this study, PdCO will be injected into the DMS and expressed selectively in dyn neurons. Following recovery, mice will learn a self-administration procedure to study reward-seeking behavior, where sucrose is delivered when mice poke their nose in the correct 1 of 2 holes, with a 5s light cue before reward delivery. Following learning, I will inhibit dyn release in a counterbalanced fashion by activating PdCO using 20 Hz pulsed 473 nm light. I anticipate that inhibiting dyn release would result in reduced sucrose consumption. Finally, I plan to use an extinction learning procedure, where dyn release is inhibited during perceived reward delivery. Here, I hypothesize that inhibition will accelerate extinction learning. By leveraging the specificity of optogenetics on various procedures, this study begins to isolate the exact mechanisms by which dyn-KOR signaling impacts reward-seeking behavior with potential insights for substance use disorder interventions.

The sensation of acute pain is fundamental to survival, indicating tissue damage that motivates an animal to engage in adaptive protective behaviors. Chronic pain, however, is pain that persists beyond the typical recovery window and serves little adaptive function. Due to the negative affective component inherent in chronic pain, it has been shown to contribute to the development of comorbid psychiatric disorders such as depression, social aggression, and social withdrawal. Our research aims to understand how acute and chronic pain affects social reward, motivation, and behavior. Our experiments used social self-administration (SSA), where pair-housed mice were placed in operant chambers and underwent voluntary lever press trials for the reward of social interaction with their cagemate. Following the acquisition of the SSA task, male and female mice then received a spared nerve injury (SNI) to induce neuropathic pain. Following recovery from the SNI surgery, experimental mice were returned to the lever press trials at different post-operative windows to simulate acute and persistent pain windows. Changes in social behavior were measured via changes in lever press frequency and interactive behavior during trials. Our data shows that male and female mice who underwent SSA trials immediately following nerve ligation maintained normal levels of social interaction. In contrast, female experimental animals that began SSA at a delayed time point, simulating persistent pain, lever pressed significantly less for a social partner compared to their respective control group. Conversely, male experimental animals at the delayed time point displayed high lever pressing behavior for a social partner. This data points towards divergent social adaptations following pain due to the sex of the experimental mice. Future experiments will further delineate these sex-specific adaptations following a traumatic injury. This research can inform social intervention as an adjunct or alternative treatment to pharmacological pain intervention and its comorbidities.

Brain-derived neurotrophic factor (BDNF) plays a critical role in neuronal function with potential implications for cognitive health, including involvement in adult neurogenesis. A decline in BDNF levels is associated with mild impairments in learning and memory. The hippocampus, known for its involvement in learning and memory processes, serves as a focal point for investigation in the brain due to its responsiveness to environmental stimuli, including exercise. There is an existing knowledge gap concerning whether running promotes an increase in BDNF levels within the hippocampus at very old ages, despite BDNF's importance in neuronal function and its potential implications for cognitive health. This study was designed to investigate whether physical exercise influences BDNF levels in the hippocampus of aged mice. Aged C57BL/6 mice were allowed access to running wheels, or locked running wheels, for three days, after which their brains were collected, post-euthanasia for neuropathology assessment. Immunohistochemistry (IHC) was performed with an anti-BDNF antibody by measuring BDNF presence, since lack of BDNF levels signifies lost neurons. QuPath digital imaging techniques were employed to provide a quantitative measure of the potential impact of running on hippocampal BDNF expression. Both the average and the variance of total distance run during voluntary wheel running decreased with age. Elevated BDNF levels were observed in the hippocampus of running mice compared to sedentary counterparts. The study provides insight into the potential impact of exercise on neurotrophic support in the aging brain. Such findings suggest a beneficial effect of exercise on neurotrophic support in the aging brain, and indicates the need for further investigations into lifestyle stratergies for promoting resilience to brain aging and cognitive decline in older adults.

Plants, like other multicellular organisms, possess a complex set of defensive measures deployed upon stimulation of the immune system. In the case of insect herbivory, these defensive measures depend upon specific pattern recognition receptors (PRRs). These PRRs enable the plant to detect herbivore associated molecular patterns (HAMPs). The Inceptin Receptor (INR) is a known herbivore receptor found in legumes, which detects Inceptin 11 (In11), a peptide fragment unique to the oral secretion of Spodoptera exigua. Beyond the plasma membrane much of the pathway from In11 detection to immune response is unknown. This research investigates possible signaling kinases downstream from INR. Receptor-Like Cytoplasmic Kinases (RLCKs) are known to mediate both specific immune and developmental signaling cascades in Arabidopsis thaliana. This suggests that they may play a role in the signaling network of herbivore induced immune responses. In this work we systematically investigate seven RLCKs of interest from three clades within the legume RLCK protein family which are upregulated by In11 treatment in the common bean (Phaseolus vulgaris). To better understand their effects, selected RLCK’s are transformed into the plant model organism Arabidopsis thaliana via Agrobacterium tumefaciens, and homozygous transgenic lines overexpressing the RLCKs are isolated. To examine the effects of over-expressing RLCKs on immunity, classic immune assays will measure their effects on Reactive Oxygen Species (ROS) and the phytohormone Ethylene production during immune activation. This research provides insights into plant immune signaling specific to caterpillar herbivory. In the future, this research may enable the intentional amplification or modulation of plant immune response to reduce losses in crops susceptible to herbivore infestation, a growing concern for food security in the near future.

RAB-2 is a small GTPase protein in the Rab family, which are known to function in dense-core vesicle (DCV) trafficking. RAB-2 functions like a molecular switch; its active form is bound to GTP, and its inactive form is bound to GDP. Activation of RAB-2 is promoted by guanine nucleotide exchange factor (GEF) proteins and intrinsic GTPase activity is dramatically expedited by GTPase-activating proteins (GAPs). The ability to switch between both forms is important to DCV maturation. RUND-1 is a known specific effector of RAB-2 that promotes the GTPase activity RAB-2, but less so than a GAP. In Caenorhabditis elegans, animals expressing RUND-1 (T237P), present a more severe locomotion defect than the null mutation. Given that RUND-1 has an effect RAB-2 GTPase activity and that the rund-1 (T237P) mutation presents a more severe defect in C. elegans, I hypothesize that the T237P mutation will either greatly promote or obstruct RAB-2 GTPase activity compared to wild-type RUND-1. To do this, I have purified RAB-2 via an intein tag and will purify the mutant protein similarly. I will use malachite green assay to measure inorganic phosphate release from GTP hydrolysis. These experiments will help help us understand the effects of the rund-1 (T237P) mutation on RAB-2 GTPase activity.

I am interested in producing Human Papillomavirus (HPV) type specific monoclonal antibodies (mAbs) from B cells derived from adolescent females and young women vaccinated with the licensed human papillomavirus vaccine, Gardasil 9, which provides protection against HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58. Fluorescently labeled pseudoviruses for these nine HPV types, as well as phenotypic markers of memory B cells are used to isolate HPV specific memory B cells via fluorescence-activated cell sorting (FACs). Gene transcripts for Immunoglobulin (Ig) heavy and corresponding light chain variable regions are amplified through Reverse Transcription Polymerase Chain Reaction (RT-PCR), cloned into their respective IgG1 backbone vectors, and subsequently transfected into eukaryotic cells to produce antibodies. HPV types [6, 11, 16, and 18] have been previously produced by our lab. I aim to expand on this work by developing antibodies against types 31, 33, 45, 52, and 58. I will characterize these mAbs for HPV binding and neutralization. Production of these well-characterized, type specific antibodies are useful for HPV studies, because they provide standards in HPV serologic assays. 

Pseudomonas aeruginosa is an opportunistic pathogen that causes chronic airway infections, a major determinant of lung inflammation and damage in cystic fibrosis (CF) patients. P.aeruginosa employs chemical signals to communicate between cells in a system called quorum sensing(QS), using several hierarchical and arranged interconnected QS circuits to regulate the expression of many genes, including those that code for virulence factors. I investigated two interconnected QS systems in P.aeruginosa, the LasR-LasI system and RhlR-RhlI system, to determine the QS hierarchy in P.aeruginosa. In the lab strain of P.aeruginosa, PAO1, the QS transcription factor LasR sits atop the hierarchy and activates transcription for many genes, including the one that encodes the QS regulator RhlR. This arrangement leads to a “hierarchy” where genes regulated by RhlR also require LasR activity. However, CF isolates have an alternate system where RhlR acts in a LasR-independent manner. We formulated the hypothesis that there may in fact be multiple architectures of quorum sensing in P.aeruginosa. The primary inquiry of this study revolves around whether clinical isolates of P.aeruginosa obtained from chronic CF infections would demonstrate continued activity of RhlR when lasR is deleted. To delete lasR, I used an allelic-exchange approach and made LasR mutants in 6 CF strains. By knocking out the lasR gene and making GFP reporter strains via electroporation, we measure RhlR and LasR activities. We found that the hierarchy as described in PAO1 is not a feature of all strains. Also, using whole-genome sequencing, we found that genes implicated in generating the hierarchy are not required in clinical strains. The findings of this project allow us to fully understand the QS hierarchy of P. aeruginosa in patients with CF, which is to determine targets for therapeutic development to combat P. aeruginosa present in patients.

The kidney is composed of thousands of filtration units called nephrons. Within each nephron lies a tuft of capillaries, the glomerulus, that filters from the blood through a filtration barrier. Over time this filtration barrier thickens, ultimately causing decreased blood filtration. A main marker of this age-related degradation are telomeres which comprise the ends of chromosomes and protect the coding DNA from degradation. If telomeres become too short, the coding region of DNA will begin to degrade. To combat this, telomere shortening signals for cells to enter a state of permanent cell cycle arrest, senescence, which prevents replication of cells with degraded DNA. Accurately quantifying telomere length will enable the development of correlations between cell lineage and structural changes within the kidney. I hypothesize that Expansion Microscopy (ExM) and quantitative-Fluorescent In-Situ Hybridization (Q-FISH) will allow me to determine the relationship between physiological changes in the filtration barrier and single-cell telomere length. ExM enables a superresolution cellular view by embedding a tissue sample in a swellable hydrogel, achieving four-fold isotropic expansion. This technique confers greater resolution of Q-FISH signal versus traditional confocal microscopy. To determine telomere length, I developed custom analysis scripts to quantify Q-FISH signal brightness. Preliminary results indicate an increased brightness of younger mice compared to their aged counterparts. Additionally, to receive a base-pair output I compared the Q-FISH signal to the signal of a DNA region of known length, Major Satellites, determining young telomeres to have an average base-pair length of 30 kb. I am validating these results in collaboration with the Miller Lab using next-generation sequencing techniques. Future work includes concurrent application of general physiology stains to identify and measure the glomerular filtration barrier physiology. Results from this method will allow for a wealth of information regarding the relationship of single-cell telomere length and glomerular structural health.

Plants' defense mechanism against herbivory and disease is integral to both natural ecological balance as well as global food supply. Induced plant responses to these threats are often triggered by specific molecules such as Herbivore Associated Molecular Patterns (HAMPS) and Pathogen Associated Molecular Patterns (PAMPS). In this project, we are refining a Luciferase Reporter Assay (LRA) and then using that assay to categorize novel HAMPS and PAMPS. This assay’s first main part is the HAMP/PAMP Receptor (HPR). By inoculating a Nicotiana benthamiana (NB) leaf with an Agrobacterium containing a Plasmid with an HPR, we are expressing specific HPRs. We are then using the second part of the LRA, a Luciferase Reporter, whose promoter is tied to an immune response related gene to measure levels of immune activity without the need for Transcriptomic Analysis. After we have infiltrated the NB leaf with the vector containing Agrobacterium and induced a response by adding a HAMP/PAMP, we are measuring Luminescence as a proxy for immune response via an imaging machine, then using R to run an analysis on the levels of immune response, helping us characterize an HPR and its signal pathway. We recently optimized time points for HPR and Luciferase imaging and have found that infiltration by Agrobacterium only takes 24 hours for sufficient plasmid integration. As such we are using this assay to run an experiment on the molecular mechanisms of an HPR in only a few days. Refining this LRA and the information we have gathered has helped shed light on the underlying mechanisms used in induced plant defense. In the future, we are planning on expressing genes of interest using this technique to seek novel activators and suppressors helping us further understand mechanisms of plant defense.

The inner ear is crucial for both hearing and balance and undergoes rapid changes during embryonic, fetal, and postnatal stages. Its complex structure poses challenges in studying auditory systems, especially the organ of Corti responsible for sound perception. Hair cells (HC’s) within the cochlea are essential for hearing, and their loss in mammals results in permanent deafness due to their inability to regenerate. We recently established Early B cell factor 1 (Ebf1) as an important factor for the development of the cochlea. Our lab’s work with conditional knockout (cKO) mouse models revealed that Ebf1 restricts sensory development within the cochlear duct. The Slc26a9-Cre Ebf1cKO model deletes on embryonic day (E) 9.5. We have seen over 2-fold increases in HC’s and their associated support cells (SCs). We have developed a tamoxifen-inducible Sox2Cre recombinase mice model for precise timing of genetic manipulation within the cochlea. Specific temporal activation of tamoxifen-inducible Sox2Cre recombinase in the cochlea will uncover critical regulatory time periods for the establishment of the sensory domain. Activation of tamoxifen-inducible Sox2Cre at embryonic day 11/12 only shows an increase in inner hair cells. These findings lead us to ask, what’s the critical window for Ebf1’s regulatory role in cochlear development? To determine the optimal window for Ebf1's regulatory role, I will activate the Cre expressed in Sox2 Ebf1-cKO mice with tamoxifen at different embryonic days (9-14) via oral gavage. Immunostaining experiments utilizing HC markers (Myo7a), inner HC markers (Vglut3), and SC markers (Sox2) will be conducted. I will quantify HC densities and cochlear length of Sox2 Ebf1-cKO and littermate control mice. Due to tamoxifen toxicity, samples will consist of embryonic day E18 specimens. Anticipated results include varying HC numbers, innervation and the presence of ectopic sensory patches. This study will offer valuable insight into the temporal dynamics of Ebf1's regulatory role.

DNA-binding proteins (DBPs) capable of targeting specific DNA sequences play key roles in genetic regulation and manipulation in both natural and synthetic contexts. Aided by advances in machine learning and protein engineering, the design of entirely new DBPs is now possible. However, early attempts yielded small, single-chain proteins that were unable to induce changes in genetic expression despite successfully binding to their target DNA sequence.  It is thought that contacts made by these early de novo DBPs are not sufficient to maintain binding in the presence of other proteins involved in DNA transcription, preventing the DBPs from altering gene expression. In contrast, many natural DBPs consist of protein complexes composed of two identical subunits called homodimers. These complexes have increased intermolecular contacts with DNA, enhancing their DNA-binding affinity. In hopes of improving the affinity of current de novo DBPs, machine learning tools like RFdiffusion, ProteinMPNN, and AlphaFold2, were invoked to engineer homodimerization domains onto these monomers. Promising designs were filtered, ordered, synthesized, and tested for efficacy as genetic inhibitors in E. coli. Homodimers that consistently and specifically bind to their target DNA sequences and induce genetic repression in E. coli have promise as tools for genetic engineering and manipulation. In future research, successful designs could also have applications in synthetic gene circuits and serve as foundations for de novo allosteric transcription factors for use as biosensors.

As our world progresses through technological advancements, much of our planet regresses as an effect of climate change, highlighting a need for underutilized resources to be brought to the forefront of industry. One avenue for transforming abundant resources into useful chemicals for generating sources like fuel is the catalytic upgrading of biomass derived molecules. However, catalysts traditionally used for these reactions are not stable to contaminants in biomass mixtures, such as water or organic acids. For biomass derived molecules to serve as precursors for biofuel and other related energy sources, more stable and efficient catalysts are needed. Our group has recently shown that a bifunctional acid–base MOF with co-localized acidic and basic sites outperforms a MOF with randomly dispersed acid–base sites for the aldol condensation reaction. To further demonstrate the importance of having the acid and base groups co-localized, I synthesized and tested three control frameworks for comparison: (1) a framework with no functionality, (2) a framework with only acidic sites, and (3) a framework with only basic sites. I then tested stability and recyclability of the bifunctional acid–base frameworks by conducting recycling experiments. I resubjected the same sample to reaction conditions for a total of 5 cycles. After each cycle, I used 1H NMR to quantify the conversion of starting material to ensure that there were no changes in catalytic activity. Lastly, I used powder X-ray diffraction (PXRD) to ensure that the catalysts maintained their crystalline structure after 5 cycles. Here I show that metal–organic frameworks (MOFs), a class of porous crystalline solids, can be used as efficient and recyclable catalysts for the aldol condensation, an important reaction for biomass conversion. Overall, this work illustrates the stability and reusability of metal organic frameworks as catalysts and thus their potential for utility in biomass upgrading reactions.

We use genes to survive and reproduce, but this means that genes can hold our survival hostage to ensure their own propagation, without providing any benefit to us. Selfish genes are a brutal, and poorly-characterized, demonstration of this concept. Instead of producing beneficial proteins, they produce nonessential proteins that prevent individuals who carry the selfish gene from successfully reproducing with non-carriers. One such example, the PEEL-1/ZEEL-1 system, is natively found in C. elegans. In this system, PEEL-1 is a toxin protein that kills cells when it is expressed without the antitoxin protein ZEEL-1. My aim is to determine the mechanism of toxicity employed by PEEL-1. AlphaFold predictions suggest that PEEL-1 contains an amphipathic helix. The amphipathic property of this region is hypothesized to play a critical role in PEEL-1 toxicity. To test this hypothesis, I am conducting a Deep Mutational Scanning (DMS) on the amphipathic helix of PEEL-1, with the goal of identifying key polar or nonpolar residues in this region that are essential to PEEL-1 toxicity. First, I generate a library of single-residue PEEL-1 mutants. Second, I transfect these constructs into HEK293T cells. I sample this initial pool of cells for sequencing, to identify which PEEL-1 mutants the pool carries, and in what proportions. Third, I induce expression, exposing each cell to the effects of the PEEL-1 mutant they carry. Only the cells expressing loss-of-function mutants survive. Now, I sequence this final surviving pool of cells, similarly to the initial pool. Mutations that drastically alter the polarity of the residue and thus break the overall amphipathic structure of the region are expected to be overrepresented in the surviving pool. This result would provide a broader understanding of the various methods of cell death in nature and provide novel insight into how animal-derived selfish genetic systems function.

It is well documented that pet cats develop age-related diseases similar to humans with chronic age-related diseases, including Alzheimer’s disease (AD). Since pet cats live in the same environment as their owners and by extension are subjected to the same environmental stressors, older pet cats are an excellent mammalian model to study therapeutic targets to slow or reverse brain aging. However, aging within the brains of pet cats is not well characterized, partly because valid reagents have not been identified. This study was designed to test several human-specific antibody reagents that identify non-neuronal cells, aging pathways, and Aβ amyloid and phosphorylated tau (pTau) seen at autopsy in brains from patients with AD. Archived brain samples, collected from pet cats at autopsy, were graciously provided by the veterinary pathology departments at University of California Davis campus and University of Pennsylvania. Immunohistochemistry staining was done to detect: 1) Microglia, a non-neuronal inflammatory reactive cell type, using an IBA1-specific marker; 2) An inflammatory pathway using an MCP-specific marker; 3) Amyloid plaques using E610, an Aβ42-specific marker; and 4) pTau fibrillary tangles using AT8, a pTau-specific marker. A digital imaging software program was used to generate a heat map to visualize staining and quantify results. It was found that brain samples from older pet cats had increased inflammation as determined by high staining intensity of microglia and MCP1. Brains from several cats showed evidence of amyloid plaques and pTau tangles. These observations suggest that the human-based reagents tested can identify analogous cell types, pathways, and pathogenic components of AD in brains from pet cats. These prototype reagents can now be used to begin the task of characterizing neuropathology in deceased pet cats donated to the Cat Alzheimer’s disease Program at the University of Washington.

Alzheimer's Disease (AD) is incredibly complex such that development of neuropathology and cognitive impairement is driven by multiple pathways. Therefore, targeting these pathways simultaneously, could provide a more effective treatment for AD compared to any single drug. Rapamycin, acarbose, and phenylbutyrate each have independent but overlapping effects on multiple pathways involved in cellular respones to pathogenic beta amyloid such as inflammation, glucose homeostasis, synaptic integrity, autophagy, and DNA damage. To test the safety and effectiveness of a cocktail of these three drugs, a proof of concept experiment was undertaken in transgenic mice carrying mutations for genes associated with early onset AD (5xFAD). These mice express neuronal amyloid plaques, a major feature of AD neuropathlogy. Transgenic and wild type mice were given either a control feed or feed containing the drug cocktail starting at 4 months of age and continued until 12 months of age. Medicated transgenic mice showed significantly less cognitive impairement in a spatial navigation learning task and reduced amyloid plaque levels in the hippocampal brain region compared to untreated transgenic mice. Immunohistochemistry will be used to identify specific biomarkers for inflammation, synaptic integrity, glucose homeostasis, autophagy, and DNA damage in the hippocampus of treated and untreated transgenic mice. Observation from this study will suggest the need to conduct additional preclincial experiments testing this specfic drug combination for a successful approach to treat Alzheimer's Disease. 

The fallopian tube function is central to several pathophysiological processes from facilitating fertilization to initiating ovarian cancer. However, existing models to study these functions are limited in relevance to the human condition due to the available conventional cell sources and culture systems. Many make use of mouse models that do not naturally develop pathologies such as ovarian cancer and lack the ability to replicate human genetic variation. Others use traditional 2D culture systems that have been shown to be poor in maintaining fallopian tube epithelium (FTE) secretory and ciliated function. Here, we propose to develop a novel in vitro model of FTE that is easy to manipulate and better mimics the in vivo environment through use of induced pluripotent stem cells (iPSC) as a cell source and a scaffold-based microfluidic culture system. We have modified an existing protocol and found that 3D culture allows for generation of organoids expressing FTE markers faster and at a higher rate. Additionally, I have developed a prototype device with a lumen and assessed cell viability and behavior of seeded immortalized FTE to establish the device. Further work can be done to elucidate drivers in FTE maturation through assessing primary FTE function in the device, and incorporation of iPSC-derived FTE cells into the device for studies on functional maturation.

The Type 2 inflammatory response occurs during intestinal parasitic helminth infection that affects over a billion people worldwide. Type 2 inflammation involves the activation of intestinal epithelial stem cells (ISCs) in the remodeling of the intestinal epithelium. ISCs differentiate into effector-like cell lineages such as goblet cells and tuft cells that lead to the production of mucus that flushes the helminth out of the intestines. However, the mechanisms that promote and regulate this epithelial remodeling and its long-term consequences remain unclear. For this project, I am analyzing the role of the bioactive lipid prostaglandin D₂ (PGD₂) in the regulation of goblet and tuft cell differentiation by creating enteroids that carry fluorescent reporters that mark ISCs. Enteroids are 3-D cell cultures formed from stem cells that model the intestinal epithelium without the presence of immune cells or other mediators; thus they allow me to hone in on the epithelial intrinsic factors that impact the immunological response in vivo. I hypothesize that PGD₂, and its receptor CRTH2, is necessary for the resolution of the Type 2 epithelial response during helminth infection. By treating enteroids with PGD₂, a PDG₂ antagonist, and inflammatory type 2 interleukins (IL) such as IL-4 and IL-13, I can analyze the role of these elements in the initiation and termination of the Type 2 epithelial response. This research is impactful because understanding the basic principles of intestinal epithelial regulation will be key to furthering the field of therapeutics and treatments for intestinal inflammatory diseases.

Myelodysplastic Syndrome/Myeloproliferative Neoplasm Unclassifiable (MDS/MPN-U) is a group of blood cancers that exhibit features of both MDS and MPN. Myeloproliferative Neoplasms (MPN) are cancers characterized by the proliferation of abnormal blood cells in the bone marrow, while Myelodysplastic Syndromes (MDS) are a group of cancers where immature blood cells in the bone marrow do not mature into healthy blood cells. Due to its overlapping features and the evolving morphology not being able to provide an accurate diagnosis, the molecular and cytogenetic abnormalities aimed at understanding the structure and properties of abnormal chromosomes could provide crucial insights for diagnosing and comprehending MDS/MPN-U. We hypothesized that the patients with MDS/MPN exhibit specific, unexplored molecular features that were not found in previous studies. We studied fourteen patients diagnosed with MDS/MPN-U overlaps and performed genetic analyses using proximity ligation sequencing (Hi-C sequencing) on Formalin-Fixed Paraffin-Embedded (FFPE) tissue specimens. Hi-C sequencing reconstructed all aberrations and rearrangements of the MDS/MPN-U genome. Cytogenetics and molecular data were collected and analyzed. New biomarkers and recurrent traits were identified. We expected Hi-C Sequencing to identify additional cytogenetics aberrations that were not detected using routine clinical cytogenetics techniques. Moreover, new insights into MDS/MPN overlap syndromes will help fill the knowledge gap in blood cancer, contributing to enhanced diagnostic accuracy, the development of treatment strategies, and improved patient care.

DNA-binding proteins contain DNA-binding domains with specific or general affinity to DNA strands and play a crucial role in multiple cellular functions, including regulation of gene expression and DNA repair. Their ability to interact with specific DNA sequences makes them attractive engineering targets for therapeutic intervention in various genetic diseases. In this project, we employed deep learning models to design novel DNA-binding proteins tailored to recognize and bind to four specific DNA sequences associated with three different genetic diseases: neurofibromatosis, sickle cell disease, and Caffey disease. These DNA sequences served as input for RFDiffusion and LigandMPNN to generate a diverse set of theoretical DNA-binding protein designs, which were then filtered with the goal of having high specificity and affinity for the target DNA sequences. These optimal designs will be expressed using yeast surface display and assayed for binding in the presence of their DNA targets, which assists in quantifying the efficacy of the deep learning models. This project aims to evaluate deep learning algorithms and to further optimize these models, as well as characterize the proteins generated with the protein design pipeline. Our designs demonstrate evidence of the feasibility of protein design through deep learning models, providing valuable data for future refinement of the algorithm. Moreover, the de novo design of DNA-binding proteins targeting DNA sequences related to genetic diseases has transformative implications for novel therapeutic interventions.

DNA-binding proteins play crucial roles in many biological processes. They can regulate gene expression by binding to specific DNA sequences and either promote or inhibit transcription, or they can serve as gene editing technology. This research focuses on the design of de-novo proteins with potential therapeutic applications in gene editing and transcriptional modulation. For our designs, we selected four short DNA sequences associated with disorders. Two of these sequences have point mutations that are associated with sickle cell anemia and hypertrophic cardiomyopathy. The other two sequences are promoter regions; one linked to the INS gene featuring a cc dinucleotide sequence that may contribute to hyperglycemia and one on the oncogene MDM2 in the MDM2p-52 pathway which has the potential to cause cancer. We then used the multi-step computational design pipeline developed at the Institute for Protein Design, which utilizes deep-learning models including RFdiffusion, LigandMPNN, and RosettaFold Nucleic Acid, to generate protein binders to the target DNA sequences. After filtering for the best theoretical designs, we would test them through yeast cell display, which visualizes the binding of the protein to the DNA. We then can analyze the binding pattern of these protein designs which vary in strength and specificity of binding to the targeted DNA sequences. Since the computational design pathway strictly filters thousands of designs to narrow our selection to the best protein designs, we expect that a few of these proteins will have significantly strong binding to their respective DNA sequences. For the designs that demonstrate strong and specific binding to the targeted sequences, we may continue to improve their structure. The success of this computational design pipeline could result in breakthroughs in protein design, and has the potential to drive medical advancement and produce treatment for genetic disorders in the future.

Cardiopulmonary bypass (CPB) is required for most cardiac surgeries. More importantly, CPB is used during heart surgeries to circulate blood out of the patient's body for surgeons to operate on the heart under optimal conditions without blood obstructing their view. However, CPB has been shown to induce systemic inflammation which can lead to complications including multiorgan dysfunction. Lack of understanding, specifically not enough knowledge of the molecular mechanisms of post-CPB inflammation has been a major obstacle to improve treatment methods. To better understand these mechanisms, we performed mRNA and Assay for Transposase-Accessible Chromatin (ATAC) sequencing on total circulating leukocytes from neonatal patients undergoing CPB. Prior data identified myeloid cells, particularly monocytes, as the major cell type driving transcriptional responses to CPB. Furthermore, Interleukin-8 (IL-8) and Tuman Necrosis Factor alpha (TNF-α) were inflammatory cytokines found to be robustly upregulated in leukocytes in patients. To further explore these findings, we performed in-vitro experiments of running THP-1 human monocytic cells to CPB-like conditions, including artificial surfaces, high shear stress, and cooling/rewarming. These experiments were collected and studied at times post shear allowing us to study and contrast how the blood is affected and recovers from CPB. Additionally, we performed drug treatment experiments with egtazic acid (EGTA) to see how calcium signaling may benefit post-CPB inflammatory response. Observing the opening and closing of chromatin of the sheared and the static blood has shown positive correlation that IL-8 and potentially IL-2 contribute to patient inflammation. We hypothesize that specifically these regions, IL-8 and IL-2, are the areas to determine where the root cause and solution are to post-CPB complications. Finally, further analysis with Topologically Associating Domains (TAD) was shown to increase chromatin density and activation in post-CPB samples in IL-8 and IL-2 genomic regions. We hope this research can reduce complications with patients going through CPB.

The mechanistic target of rapamycin, mTOR, functions in the mTORC1 complex with another protein called raptor as a master regulator of eukaryotic cellular metabolism thereby regulating cell growth including from cancer, cell death including after stroke, inflammation, and aging. In a forward genetic screen for hypoxia resistant mutants, the Crowder lab recently identified a missense mutation in the daf-15 gene, which encodes C. elegans raptor. The mutation produces a heat-sensitive reduction of raptor function, hereafter referred to as daf-15(rf). At 20°C, daf-15(rf) is normally hypoxic sensitive, at 22°C very hypoxia resistant, and at 25°C incapable of normal development. Raptor negatively regulates autophagy, a mechanism for breakdown and recycling of proteins and organelles. Activation of autophagy has been found to promote hypoxic survival in C. elegans and higher organisms. Thus, we hypothesized that activation of autophagy was responsible for the hypoxia resistance of our daf-15(rf) mutant. To test this hypothesis, we first asked whether we could detect increased autophagy using fluorescently-tagged autophagy proteins at 22°C in daf-15(rf) but saw no effect compared to wild type. Next, we asked whether a C. elegans transcription factor, HLH-30, that promotes expression of autophagy proteins was activated by daf-15(rf) and found activation at 25°C but not at 22°C. Finally, we tested whether proteins essential for autophagy were also necessary for the hypoxia resistance of daf-15(rf). By generating double mutant strains, we showed that animals with daf-15(rf) but without essential autophagy proteins were still hypoxia resistant. Thus, we conclude that C. elegans raptor regulates hypoxic sensitivity by an autophagy-independent mechanism. These findings demonstrate that raptor can control hypoxic cellular injury by mechanisms distinct from autophagy. Such mechanisms, if identified, could be targeted for treatment of cancer, stroke, and other diseases where hypoxia plays a role.

Over the past few years, patients have been identified with debilitating phenotypes due to mutations in MSTO1, a nuclear gene. These patients often have distal muscle loss and weakness leaving patients incapable of walking but to date there is no known treatment. One barrier to progress is that virtually nothing is known about MSTO1 function, making the development of therapeutics for these patients extremely challenging. The goal of this project is to use an unbiased approach to discover functions of MSTO1. To do this, I will find genetic interactors utilizing yeast to perform an unbiased screen. Yeast DML1 is the homolog to MSTO1 and is required to keep the yeast cells alive. This screen will identify genes in the yeast genome that support survival of cells lacking DML1 when the gene is overexpressed. We utilize an auxin-degron system that targets DML1 for degradation when the yeast are grown with auxin. To find genes from the yeast genome that keep the cells alive when DML1 is degraded, I express random fragments of genomic DNA. Those genes must, therefore, be linked to DML1 function in some way, thus providing insight into what MSTO1 does, how it works, and how to help MSTO1 defective patients. I have obtained hundreds of yeast colonies that survive without DML1 when other genes are overexpressed. Currently, I am extracting these overexpressed DNAs to determine the gene(s). This work is an essential step toward fully understanding MSTO1 function in cells and we plan to characterize these connections in yeast and human cells.

Epigenetic factors, including histone marks, change the patterns of gene expression without altering the DNA sequence. Variations in such marks are known to account for the ability of stem cells to differentiate into various cell types, but a preliminary experiment conducted by a former member of the Vaughan Group, Dr. Marcus Woodworth, has revealed that even in a phenotypically homogeneous, terminally differentiated cell population, the presence of H3K27me3, a repressive histone mark, varies on the HOXC gene of human retinal pigment epithelium (RPE1) cells at single-cell, single-loci level. My role is to evaluate the two possible origins of such heterogeneity: inheritance (histone mark varies due to events that happened during differentiation, or the random drift after differentiation, and the variations are kept within each lineage of cells), and multiple mark co-repression (one histone mark varies, but summing its effect with another histone mark that perform a similar function lead to the observed functional homogeneity), and to validate that such pattern exists among a broader range of genes. To achieve these ends, I profile selected histone marks (H3K27me3 and H3K9me3) on genes that experience different types of regulations during differentiation (HOXC, GAPDH and SIX6), using imaging-based methods, including the time-lapse imaging of live cells to map out lineages, and expansion microscopy (ExM) to capture fluorescently labeled histone marks at single-loci level. If the hypothesized origins are true, a significant difference in the number of histone mark clusters around the genes of interest would be observed between cells of different lineages, and complementary variation patterns would be observed between H3K27me3 and H3K9me3. The study reveals the nuances of histone mark dynamics on the single-cell, single-loci level, and optimizes an imaging-based method that has the potential for multiplexing at high spatial resolution, thereby providing a powerful tool for further studies on epigenetics.

Neuroscience research relies heavily on cell counting to assess brain injury and evaluate neuroprotective treatments. While manual methods (i.e. hand counting) have been used traditionally, automated programs offer the potential for standardized and error-free data analysis. In the context of studying hypoxic-ischemic encephalopathy (HIE), a prevalent brain injury in infants, we aimed to assess the accuracy of an automated cell counting program in an in vitro slice culture brain injury model of injury and treatment. Code templates from ImageJ/Fiji were taken and modified using ChatGPT, and other snippets were used and modified from online forums like GitHub. The program searches through folders/subfolders for images, converts them into a binary image based on fluorescence threshold data (used to stain cell nuclei), applies the ImageJ function “watershed” that breaks larger groups into smaller groups, runs the “analyze particles” function which outputs a total cell count based on the size and circularity of the cells, and then saves the final image. Fluorescence threshold, cellular size, and circularity values were determined before data collection by adjusting the values to best fit the final image of a random slice. The settings were then kept consistent within studies. Preliminary results show the program’s high accuracy and precision, with consistent results across caffeine and Azithromycin treatments in our in vitro injury model. Despite yielding higher counts than manual methods, the program remained consistent across models. Validating this automated method represents a significant advancement in research methodology. These programs offer standardized data collection, error elimination, and faster analysis compared to manual counting, potentially saving time and resources for labs. Current limitations in our research involve differentiating between healthy and dead or dying cells, which would be an important future step for automated cell counting.

Mitochondria are the main oxygen consumers in eukaryotic cells and as such are the primary organelles affected by oxygen deprivation, hypoxia. Hypoxia alters the size and shape of mitochondria, called the mitochondrial dynamics, but their role in hypoxic cell death is unknown. The Crowder lab has recently discovered that a mutation in the Mechanistic Target of Rapamycin Complex One (mTORC1) protein Raptor confers hypoxia resistance in the nematode C. elegans. mTORC1 is a master regulator of metabolism and is known to affect certain aspects of mitochondrial biology. Given these two facts, we tested the hypothesis that the hypoxia resistance of the C. elegans Raptor mutant is from alterations of mitochondrial dynamics. First, I showed that hypoxia induces small, rounded mitochondria in C. elegans caused from mitochondrial fission. Second consistent with the hypothesis, I showed that the mitochondria appear to have more normal morphology before and after hypoxia in the Raptor mutant. However, not consistent with the hypothesis, a C. elegans mutant with excess mitochondrial fission was not hypersensitive to hypoxia. Then combining the hyper fission mutant with the Raptor mutant did not diminish the hypoxia resistance produced by reduced Raptor function. Thus, our data demonstrates abrogating mitochondrial fission is not necessary for the hypoxia resistance produced by the Raptor mutant and leads us to reject our hypothesis. By exploring the interaction of mitochondrial fusion and fission with Raptor, we are beginning to understand how these important organelle and metabolic regulators combine to control hypoxic cell death.

This interdisciplinary project has the goal of investigating the interplay between behavior and biology using the context of thermoregulation in an invertebrate animal model system. Specifically, our approach involves the design, construction, and implementation of an ethoscope apparatus that examines behavioral responses associated with exposure to ethanol vapor (EtOHvap) in Drosophila. The methodology involves integrating a customized EtOHvap delivery and removal system into an ethoscope apparatus into which we incorporate a thermal gradient within the chamber. This customized ethoscope allows for the assessment of Drosophila behavioral responses prior to, during, and after EtOHvap exposure. Analysis of these data will focus on a comparison of the behavioral responses and how they reflect the effects of acute EtOHvap exposure in the form of temperature-preference-based behavior. Equipped with a Raspberry Pi and HD camera, the ethoscope apparatus captures and records Drosophila movements with software using OpenCV-Python to analyze the resting positions along the thermal gradient, and then compares them with the post-exposure resting positions. By working with Drosophila, a well-studied animal model, our design allows us to rapidly identify and examine behavioral responses as potential indicators of thermoregulatory responses to EtOHvap and forms the basis of future directions of research that utilize behavioral responses to understand underlying body temperature physiology and homeostasis. This research intersects with other works that have reported on the effects of ethanol exposure in other contexts and different model systems, including vertebrates. Thus, findings from this research not only inform this specific investigation but also have implications for understanding differences in how different animals regulate body temperature through behavior.

The seeds and fruits of all flowering plants derive from female reproductive structures called pistils. At the tip of a pistil is an area called the stigma that is the site of pollen reception, making the structure of stigmas critical for ensuring plants are pollinated and produce seeds. In the genus Thalictrum, stigmas vary widely: short stigmas are present in insect-pollinated Thalictrum species to facilitate closer contact with visiting insects and ensure successful pollen deposition, while longer, more feathery stigmas have evolved from these to take advantage of wind for pollination. However, the evolutionary and genetic basis for this transition is not known. I aim to explore the genetic mechanisms that facilitate the transition from insect-mediated to wind-mediated pollination using Thalictrum thalictroides as a model species. Specifically, I am testing the function of two candidate genes in T. thalictroides stigma development, STYLISH1 and STYLISH2 (STY1/2), which are known to be necessary for stigma development in other plant species. To do so, STY1/2 genes have been silenced and overexpressed by virus-induced gene silencing and virus-mediated overexpression. I collected and imaged floral tissue from transgenic plants and validated them by qPCR. The effects of silencing and overexpressing STY1/2 are then determined using scanning electron microscopy to visualize cellular changes to the pistil and stigma. I predict shortened or loss of stigmas in silenced flowers and elongated stigmas in overexpressed flowers, which would support my hypothesis that STY1/2 are necessary for stigma formation in T. thalictroides. Not only do these findings enhance our knowledge of plant adaptation to environmental changes, but they also create implications for efforts toward conservation and sustainable agriculture, offering potential insights into the prospect of engineering wind-pollinated plants in a changing climate with dwindling pollinator populations.

DNA-Binding Proteins (DBPs) hold a strong affinity to interact with the major grooves of DNA for the purposes of transcription, translation, and repair. Although DBPs are found in nature, their specificity is difficult to predict and their production expends excessive resources. Therefore, our project’s goal is to efficiently generate DBPs that allow us to enable exact processes to occur. This is promising for future use in treating genetic diseases. In our research, we studied point mutations in the hexosaminidase subunit alpha (HEXA) gene and adenosine deaminase (ADA) gene, which can lead to Tay-Sachs disease (TSD) and Severe Combined Immunodeficiency (SCID), respectively. TSD is a rare genetic disease that affects infants and ultimately leads to brain damage, resulting in these children not making it to grade school age. Similarly, SCID is genetic, where children lack a strong immune system. This increases their susceptibility to infections, especially during their first year of life. Targeting the HEXA and ADA genes, we first developed designs utilizing computational software like RosettaFold, RFdiffusion, x3DNA, and LigandMPNN, followed by rigorous filtering via RosettaFold Nucleic Acid. Afterwards, we tested the final designs using yeast cultures and fluorescence-activated cell sorting (FACS) in the laboratory to determine which bind best to their generated DNA template sequences. Overall, we expect to find a few specific DBPs that bind effectively as predicted during the computational pipeline. These successful designs can be utilized as genome-editing proteins, correcting their target DNA sequences and restoring normal function.

Pediatric high-grade gliomas (pHGG) are highly aggressive brain tumors with limited therapies and extremely poor clinical outcomes. pHGG and many other cancers are often driven by genetic fusion events, which produce oncogenic fusion proteins such as TPM3-NTRK1. NTRK1 regulates signaling pathways promoting cell survival and differentiation. In cancer, NTRK fusions lead to constitutive activation of these pathways, resulting in uncontrolled cell growth. The purpose of my project is to validate TPM3-NTRK1 as a potential drug target for a novel therapeutic strategy known as targeted protein degradation and to clarify the role of TPM3-NTRK1 in cancer signaling. I hypothesize that degrading the TPM3-NTRK1 protein will decrease the activity of downstream signaling pathways that lead to pHGG. To model pharmacological degradation of TPM3-NTRK1, I applied the degradation tag (dTAG) system to rapidly and reversibly degrade a target protein fused with an FKBP12^F36V-tag. In this approach, dTAG molecules bind to the FKBP12^F36V-tag and recruit an E3 ubiquitin ligase to ubiquitinate the target protein, leading to its degradation by the proteasome. In my project, I used Gateway cloning to generate a lentiviral expression plasmid containing FKBP12^F36V-TPM3-NTRK1. I then produced lentiviruses, which I used to transduce NIH/3T3 cells and express FKBP12^F36V-TPM3-NTRK1. To test whether the FKBP12^F36V-TPM3-NTRK1 protein can be successfully degraded, I plan to treat the cells with dTAG molecules and use Western blotting at various time points to evaluate levels of FKBP12^F36V-TPM3-NTRK1 and known proteins involved in downstream signaling pathways, including pERK and pAKT. I expect that samples treated with dTAG molecules will show significantly decreased FKBP12^F36V-TPM3-NTRK1, pERK and pAKT. Importantly, these results will justify the development of small-molecule degraders to target TPM3-NTRK1, a promising therapeutic approach for pHGG and other hard-to-treat cancers. Additionally, a cell line expressing degradable TPM3-NTRK1 will serve as a powerful model system to further explore its role in cancer.

Alzheimer's disease (AD) is a neurodegenerative age-related disease characterized by the presence of amyloid-beta aggregates and hyperphosphorylated tau tangles. It has been well documented that cognitive decline and changes in age-related pathways are associated with disease progression. Mitochondria play an important role in degradation of amyloid protein through a mitochondrial protein-mediated quality control system. This pathway can break down with increasing age and lead to the overwhelming presence of amyloid, disrupting normal mitochondrial activity. This damage leads to the formation of more Aβ plaques and neuroinflammation, contributing to the pathogenesis of AD. Mitochondrial regulators may be potential therapeutic drug targets but models are needed to help identify and characterize them. In this regard, an Adeno-Associated-Viral (AAV) vector was used to induce AD protein expression in the brains of old mice. 40 Male and 40 Females mice aged 24 months were infected with either the AAV-AD or AAV-SHAM vector and given 3 months for expression of the proteins to build. Mice were euthanized and brain tissue collected into formalin, with the hippocampus cut into slides for immunohistochemistry (IHC). Data generated from these mice has shown trends in decreased synaptic integrity, increased inflammation and DNA damage associated with expression of the vector proteins. Utilizing the same model, this experiment aims to understand how expression of the AAV-AD proteins may be associated with known roles of mitochondria and characterized pathways in the early stages of AD. IHC was performed using antibodies specific for PITRM1, a mitochondria protein degradation regulator, and PINK1, responsible for mitochondrial-mediated cell death (mitophagy). Imaging software “ImageJ” will be used for quantitative analysis of the stains. This study will help clarify an association between varying levels of AD protein expression and mitochondrial regulation, providing valuable information for enhancing therapies aimed at preventing the progression of early stage AD.

The causative agent of malaria, Plasmodium spp., generated 608,000 deaths worldwide in 2022 according to the World Health Organization and disproportionately threatens endemic areas of Africa. Plasmodium sporozoites infect the host by entering the bloodstream through the skin following bites by female Anopheles mosquitoes. From there, sporozoites migrate to the liver and infect hepatocytes. A single sporozoite-infected hepatocyte is capable of producing thousands of merozoites, which go on to enter the bloodstream. Complete elimination of infected hepatocytes is necessary to achieve sterile protection. In order to observe adaptive and innate immune cell localization towards infected hepatocytes, we applied fluorescence microscopy on livers in a BALB/c rodent model of malaria. Naïve unvaccinated mice were infected with sporozoites of Plasmodium yoelii, a rodent malaria parasite. Two important cell populations are recruited to infected hepatocytes. The first are tissue resident memory CD8+ T cells (Trm), which are crucial in pre-erythrocytic protection. The second are Kupffer cells, which are specialized liver macrophages. To measure these adaptive and innate cell populations, respectively, we applied fluorescently-labeled antibodies to mark the parasite as well as Trms and Kupffer cells. After staining the collected liver tissue and imaging with a widefield fluorescent microscope, we visualized recruitment and measured immune cell proximity quantitatively within a region of interest of the area surrounding infected hepatocytes using microscopy imaging analysis software. This method will be used to test the hypothesis that Trms and Kupffer cells are induced following sporozoite challenge in the rodent malaria model.

One of the most common types of vaccines used today are subunit vaccines. Subunit vaccines consist of an antigen that triggers the adaptive immune system to create antibodies but also require a separately added adjuvant, which is a substance that induces longer-term immunity by stimulating the immune system to pay attention to the antigen. Current adjuvants are non-specific - often things like oil-water emulsions that irritate the immune system and cause inflammation in unknown ways to draw attention to the antigen. This project aims to create a more specific adjuvant by directly stimulating B cells. In order for B cells to replicate antibodies, they need a primary signal from the antigen and a secondary signal that certain ligands on T-cells can initiate. We decided to investigate whether CD40Ligand (CD40L), an immune protein present on T cells that works to signal B cells to either replicate or create antibodies, could be used to achieve this goal. The idea behind the project is to co-display CD40L with antigen on the nanoparticle in hopes of creating a more specific adjuvant. We designed 10 different versions of this nanoparticle, where we tested two versions of CD40L, the placement of CD40L, and the linker length between CD40L and the nanoparticle surface. Our preliminary results also show that our cages retain their ability to bind both antibodies and CD40 as well as activate NFkB transcription - a proxy for B cell activation. We expect CD40L-displaying nanoparticles will promote B-cell proliferation to a greater extent than the nanoparticle vaccine displaying only hemagglutinin (flu) antigen accompanied with an adjuvant like Addavax. Ultimately, we hope to examine how co-display of CD40L with antigen will change the quality of immune response and memory in-vivo in comparison to currently used vaccine adjuvants, and begin testing in-vivo in the coming quarters.

Glomeruli are the basic filtration unit of the kidney. The current understanding of its physiology is limited by the partial or 2D analysis of its structural components. The Vaughan Group uses optical super-resolution microscopy in combination with advanced chemical labeling techniques and powerful data analysis approaches to perform high-resolution 3D reconstruction of the whole mouse glomeruli. Overall, the work has the potential to provide a novel understanding of the glomerular structures and how they are altered in aged and diseased conditions. The labeling of the overall morphology of the glomeruli is achieved by chemically labeling the distribution of abundant macromolecules (carbohydrates, amine, and DNA) using Fluorescence Labeling of Abundant Reactive Entities (FLARE). Though we could visualize the general physiology of the sample with FLARE, incorporating specific targeting of molecules with FLARE is still challenging. My role is to optimize the FLARE protocol to add the capability of labeling the distribution of specific molecules using immunolabeling. The most challenging part is that all the fluorophores labeled prior to FLARE will be bleached out by the strong oxidation step while labeling carbohydrates. I am focusing on exploring possible workarounds to incorporate immunostaining with FLARE. The only way to bypass the bleaching fluorophores is to label dyes after the FLARE. However, the FLARE involves the gelation part, and the gel makes antibodies which are linked to fluorophores hard to get into the sample. So, instead of using regular secondary antibodies, I use biotin and then link to the streptavidin dye, which is smaller in size and easier to enter the sample. With this optimization working, we could incorporate whatever target of interest with high resolution on top of three general stains provided by FLARE, giving us an extra degree of information for our 3D reconstructions of glomeruli.

Plants utilize cell surface protein receptors to recognize insect herbivory through the detection of Herbivore Associated Molecular Patterns (HAMPs) Following the detection of HAMPs, plants initiate specific immune responses, often measured by the increased production of the hormone ethylene gas and by Reactive Oxygen Species (ROS). The Inceptin Receptor (INR), which is specific to legume plants, recognizes the HAMP Inceptin11 (In11). The binding of In11 to INR initiates a signaling cascade, leading to an immune response. However, the signaling mechanism activated by INR is unknown. The Receptor-Like Cytoplasmic Kinase (RLCK) gene Herbivore Induced Kinase 1 (HIK1) is upregulated by In11 treatment in the bean species Phaseolus vulgaris. The goal of this research is to determine if HIK1 and other RLCKs are downstream proteins required for INR signaling. Because of the genetic intractability of P. vulgaris, I transform Arabidopsis thaliana with RLCK genes using Agrobacterium tumefaciens infiltration. Isolated genetic lines are then used to analyze the effect each RLCK has on immune signaling. Transgenic plants are treated with bacterial associated molecular patterns to trigger an immune response, then tissue samples of the leaves are measured for ROS and ethylene gas production. Results are then compared with ROS and ethylene gas production of wildtype plants. If the studied RLCKs are involved in downstream INR signaling, the transgenic plants will have increased ROS and ethylene gas production. I anticipate HIK1 to have the strongest increase in ROS and ethylene gas production due to the upregulation of HIK1 after In11 treatment in P. vulgaris. Understanding the INR signaling pathway is vital for engineering of plants that are resistant to insect herbivory without the use of pesticides.

H3K9me3 heterochromatin regulates gene silencing and plays key roles in genome stability and cellular identity. During cell reprogramming, H3K9me3 has been shown to block transcription factor binding and changes to cell identity. Recently, enhancer of rudimentary homolog (ERH) was identified as a master regulator of H3K9me3 and a repressor of reprogramming in humans. However, the roles that specific ERH protein amino acids play in its function remain unknown. We hypothesize that elucidating the role of ERH and identifying pivotal amino acids will enhance our understanding of its molecular mechanisms. We are targeting amino acids which are evolutionarily conserved as these are more likely to be important for ERH function. We have shown that the ERH knockdown phenotype of low proliferation and decreased H3K9me3 can be rescued by exogenously expressing a wild-type copy of ERH. Our approach has involved generating mutant variants of ERH, introducing them into cells where the endogenous ERH is depleted and then assessing whether they rescue the phenotype. To efficiently assess many amino acid substitutions in the ERH protein simultaneously we are developing a multiplexed alanine scanning approach with Nanopore readout in a functional screen. In this approach, multiple ERH mutants will be simultaneously introduced and integrated into the genomes of a population of cells with the endogenous ERH depleted. Mutant forms of ERH which do not rescue the proliferation phenotype are expected to decrease in abundance over subsequent proliferation. The ratio of exogenous ERH sequences will be assessed upon initial introduction and after 2 weeks by targeted long read DNA sequencing. Identifying which amino acids are important for ERH function will allow us to design further experiments, investigating their role in localization, protein-protein interactions and H3K9me3 regulation. Understanding the regulation of H3K9me3 heterochromatin will be essential to improve cell reprogramming, enabling autologous transplantation and regenerative therapies.

Bees collect a variety of pollen as a protein source to feed their brood. Any decline in the concentration of protein in pollen can be detrimental to the survival of younger bees, and any environmental factors that decrease the amount of protein in pollen can therefore have downstream effects on the overall health of honeybee colonies. As part of a longer longitudinal study, this study explored the difference in protein concentration in pollen collected in 2022 and 2023. This short term study is part of a larger project in which we seek to discover if there is a correlation between the changing climate and the changes in pollen protein concentration. We used a Bradford Assay to determine the protein concentration in pollen collected from hives at Edmonds College during equivalent time periods in 2022 and 2023 . We also subjected the pollen samples to microscopic analysis following an acetolysis procedure, in order to view the pollen's characteristics and floral type. Our results showed a statistically significant drop in protein concentration (two sample t-test, p< 0.05) from 2022 to 2023. As the climate continues to rapidly change around us there could be unforeseen impacts on bee health and populations. As this experimental project continues throughout the years, we hope to understand the full impact of climate change on the concentration of protein in pollen.

Investigating the mechanisms behind asymmetric cell division (ACD) not only provides insight into an integral aspect of development in organisms such as C. elegans, D. melanogaster, and in human stem cells, but it also has the potential to elucidate the cellular functions that result in disease processes like cancer. ACD describes the cellular activity that results in a singular parent cell dividing into two daughter cells with two distinctive molecular compositions, and, in turn, fates. Drosophila melanogaster neural stem cells, called neuroblasts, are intrinsically polarized and divide asymmetrically. This process involves an uneven distribution of cell-fate-determining proteins that eventually designate the function of the daughter cell. Because of this polarity establishment and orientation, these daughter cells can have highly specialized functions. My work uses Drosophila melanogaster neuroblasts to screen for novel mutations responsible for compromising ACD. I'm specifically interested in finding genes that are involved in the generation of cell size asymmetry. Finding these mutations is imperative because, by doing so, we can identify critical genes involved in this process of interest. To that end, I knocked-down candidate genes with RNAi in neuroblasts and assayed the localization dynamics of myosin and microtubules– two critical components of the ACD and symmetric division process. Through confocal, live-cell imaging, I then examined the dynamics of these cellular proteins as they relate to wild-type ACD to see if my chosen candidates were responsible for this process. I anticipate finding novel cytoskeleton regulators that are important for establishing cell size asymmetry. In the future, finding the genes behind ACD will provide great insights into the mechanisms behind both development and disease in addition to the field of stem cell biology overall.

This study explores the adaptive dynamics of Pseudomonas aeruginosa (P. aeruginosa), a bacterium that often plagues cystic fibrosis patients. In P. aeruginosa, many genes are regulated through quorum sensing (QS), a form of cell-to-cell communication. In wild-type P. aeruginosa, strain PAO1, LasR acts as a QS regulator, controlling several genes, including those encoding proteases, as well as two other QS regulators, RhlR and PqsR. QS-regulated proteases are secreted by P. aeruginosa into the environment, benefiting the whole population– a cooperative process. When grown in conditions requiring proteases for growth, such as on casein, mutated P. aeruginosa emerge that are capable of using proteases without producing any themselves. While disadvantageous to the community, these “cheaters” experience a fitness advantage. Over time, the proportion of cheaters increase until the population cannot support itself and collapses. Related to QS is the transcription factor MexT, that, when active, down-regulates RhlR. Another transcription factor, MexS, down-regulates MexT expression. In PAO1, MexS is inactive, resulting in constitutively expressed MexT. I was interested in how variants of MexT and MexS that alter expression of QS regulators might influence cooperative behaviors in P. aeruginosa. I compared survival and function of wild type PAO1 to two engineered strains, one with restored MexS function (MexS+) and one with MexT deleted (ΔMexT). I passaged these strains in casein broth for 30 days, evaluating every five days for the emergence of protease-negative mutations. I am also analyzing the genetic composition of each population, focusing on categorizing lasR and a gene involved in peptide transport, psdR. Preliminary results show fewer mutations in psdR than expected in all strains, and fewer mutations in lasR in the ∆mexT strain. Continued exploration of the complex QS and gene regulatory pathways in P. aeruginosa may point to mechanisms of virulence and to potential targets for antibiotic therapies.

Cells' ability to efficiently replicate their genomes is essential for regulating chromosomal division and maintaining chromosome integrity.  Defects in any of these cellular processes may cause genomic instability, potentially leading to cancer.  The Chaos3 allele in the yeast Saccharomyces cerevisiae is a single base pair change causing an amino acid substitution in the Mcm4 protein.  Mcm4, a component of the replicative helicase, is recruited to replication origins to unwind double stranded DNA and initiate replication.  Chaos3 is in a region of MCM4 that is highly conserved across eukaryotes; while mutations in conserved regions are generally non-viable, Chaos3 is a viable allele that causes genomic instability, leading to elevated cancer rates in mice.  In S. cerevisiae, Chaos3 decreases early firing of the autonomously replicating sequences (ARS) where DNA replication begins.  Chaos3 does not affect all early firing ARSs in the genome; rather, a large proportion of origins near centromeres, thereby delaying replication of those centromeres.  Essential for chromosome segregation, the centromere is the location where spindle fibers attach to pull apart sister chromatids during cell division.  I hypothesize that this delay in centromere replication results in chromosomal instability, including the loss of a chromosome.  I am using CRISPR guided cutting directed by a customizable guide RNA to replace centromeric adjacent ARS510, that has decreased firing levels in Chaos3, with unaffected, early firing ARS305, to see if firing levels in the mutants are affected based on ARS chromosome location (i.e., proximity to a centromere) or ARS sequence.  If replacing ARS510 with ARS305 restores early origin firing in this region this will confirm the Chaos3 mutation affects specific ARS sequences rather than ARS location on the chromosome.  Furthermore, if centromere replication delays are the cause of genomic instability in Chaos3, this ARS replacement should rescue chromosome loss.

mTOR, the mechanistic target of rapamycin, is a serine/threonine kinase that regulates protein synthesis, cell growth, and metabolism in response to nutrients and energy in most eukaryotes. mTOR consists of two distinct complexes, mTORC1 and mTORC2. These complexes can be further divided into three components: mTORC1 consists of mTOR, Raptor, and mLST8, and mTORC2 consists of mTOR, Rictor, and mLST8. mTORC1 is critical in metazoan development and has been implicated in aging, cancer, diabetes, cardiovascular disease, and hypoxia. Previously, the Crowder lab conducted a mutant screen in C. elegans for hypoxia resistant mutations, and identified a missense reduction of function mutation in the daf-15 gene, the C.elegan ortholog of Raptor. A unique feature of this mutation is that the function of Raptor can be turned on and off by varying temperature. It has normal hypoxia resistance at 20 degrees, increased resistance between 21-22, and developmentally arrests at 25 degrees. I and the other authors made use of this conditional developmental arrest phenotype to screen for genetic suppressors. Using genetic mapping, sequencing, and complementation testing, we have identified multiple mutations in three different genes responsible for restoring Raptor function. One of the genes identified in this manner was rnf-126. Results show mutations in rnf-126 suppress the Raptor mutation. A null mutation in rnf-126 similarly suppressed the Raptor mutation. Previous work has implicated mammalian rnf-126 in degradation of the mTORC1 complex in cancer cells, suggesting that reduced levels of daf-15 may produce hypoxia resistance. We tested this hypothesis using auxin-mediated degradation of daf-15, finding that auxin-treated animals are hypoxia resistant. Current work by myself and others will further investigate how rnf-126 controls Raptor function and hypoxia sensitivity. Elaborating the function of this gene will define novel mechanisms whereby Raptor and mTORC1 controls metabolism, hypoxic injury, and development.

Pembrolizumab (brand name KEYTRUDA), is a monoclonal antibody therapy classified as an immune checkpoint inhibitor, a type of immunotherapy that works by facilitating the detection of cancer cells by T cells. Specifically, it blocks the PD-1 pathway that cancer cells use to suppress and evade the immune system. With neoadjuvant-adjuvant treatment, patients receive systemic therapy both before and after the surgical resection of their tumor(s). There is limited research evaluating the association between tumor responses (changes in tumor measurements) and survival time with neoadjuvant anti-PD-1 treatment. In this study, I analyzed the association between tumor measurements and event-free and overall survival in a clinical trial of patients receiving neoadjuvant-adjuvant anti-PD1-therapy. Event-free and overall survival were estimated using the Kaplan-Meier method, and associations with RECIST tumor size were evaluated using martingale residual plots and Cox proportional hazards regression models. In this study, I found significant associations between event-free survival and baseline tumor burden and changes (quantitative and categorized) in tumor size between baseline and the end of neoadjuvant therapy. Increases in tumor size were associated with a lower 2-year event-free survival. Larger values of baseline tumor burden were significantly associated with event-free survival but were not significantly associated with overall survival. The findings from this study may serve to better inform clinicians on making prognoses for patients on anti-PD-1 therapy and indicate a need for more research to be done on associations between tumor size and overall survival outcomes.

Alzheimer’s Disease (AD) is a progressive brain disorder that debilitates memory, learning, and decision-making. Early-stage AD represents the initial phase where individuals are still able to function independently, but with increasing age, their condition steadily progresses to dementia and loss of independence. Because a significant number of the aging population is affected by AD, understanding the neuroinflammatory processes would help develop more effective strategies for treatment. Examining markers such as MCP-1 and TNF-alpha, known to be associated with inflammatory response, will help identify the modulatory processes that lead to mild cognitive impairment associated with early-stage AD. Subsequently, higher levels of inflammation markers within the brain leads to mild cognitive impairment. This research study involved 40 C57BL/6 mice, 20 males and 20 females (21 months old), retro-orbitally infected with 80 µL of neurotrophic AAV-AD vector or AAV-Sham for a duration of 2 months before humane euthanasia. Brains were collected, and specific regions were examined by immunohistochemistry (IHC) and digital imaging to assess the expression levels and distribution of the inflammation markers. Preliminary observations showed that hippocampal regions of the brain from mice with early-stage AD had higher staining intensity for MCP-1and TNF-alpha compared to respective areas in Sham mice, suggesting increased inflammation is a very early lesion that develops in the presence of AD pathogenic components that might be controlled by anti-inflammatory drugs. The preliminary data suggests that the characteristics of AD manifest in part due to the neuroinflammatory response of brain factors that change with onset AD.

Mutations in phosphodiesterase 6 (PDE6) underlie photoreceptor degeneration through cyclic guanosine monophosphate (cGMP) accumulation, triggering a series of down-stream processes, which eventually kill photoreceptors. We hypothesize that knocking out inosine monophosphate dehydrogenase 1 (IMPDH1), the rate-limiting enzyme in de novo guanine synthesis, will rescue cell death caused by PDE6 mutations. Supporting evidence from a mouse mutant model (rd10) suggests that inhibiting IMPDH1 pharmacologically delays photoreceptor degeneration (Yang, 2020). Our procedure for this experiment is as follows. Fish heterozygous for impdh1a and pde6c mutations are mated to produce fish that are double homozygotes. Fish homozygous for mutations in both the cone-specific pde6c and the impdh1a genes are genotyped and embedded for histological analysis of the retina. Histology is examined on days 3,5, and 7 post-fertilization (dpf) for cone degeneration. To date, we have genotyped our two mutant lines. Normally, pde6c^-/- fish have severe cone photoreceptor degeneration at 5 dpf and impdh1a^-/- fish show no signs of photoreceptor degeneration even as adults. If degeneration is rescued, the double knockout larvae should retain similar photoreceptor nuclei counts to wildtype fish at all time points. Demonstrating that IMPDH1 inhibition rescues PDE6 deficiency would provide proof-of-concept for the therapeutic potential of IMPDH1 targeted inhibition for the treatment of photoreceptor degeneration due to cGMP imbalance.

Lewy Bodies (LB) are É‘-synuclein protein aggregates that form when the protein is falsely over-expressed. This inappropriate cell behavior is a symptom of neurodegenerative ailments including Parkinson’s Disease. GATA-1 is a transcription factor that binds to a conserved region in the SNCA gene and promotes transcription of É‘-synuclein mRNA, which could be a contributing factor to disease presentation. DNA binding proteins serve a host of functions, primarily pertaining to regulation of gene expression. Through the Institute of Protein Design and their computational design pipeline, I designed proteins that bind to the GATA-1 binding site in the SNCA gene with the intention of inhibiting GATA-1 and hyper-expression observed in Parkinson’s patients. First I modeled the DNA target sequence using Chemnet software to best approximate the B form DNA structure that the final optimized proteins bind to. RoseTTAFold Nucleic Acid Diffusion generates protein backbones that have strong shape complementarity for the grooves of the target DNA. I then used LigandMPNN software to produce amino acid sequences for each of the backbone structures generated in the previous step. Designs are refined and diversified before the most plausible binders are selected via statistical metrics, AlphaFold2, and RoseTTAFold. I yielded roughly 5000 DNA binder models that were ordered as DNA libraries for high-throughput testing. Proteins are then expressed using yeast surface display and sorted via flow cytometry, where fluorescence indicates a binding event between the protein and DNA target. I had a robust sample of design outputs and intend on analyzing the successful protein models to quantify the importance of different structural features in DNA binding. Successful binders not only concurrently optimize and evaluate the efficacy of these deep learning models, but demonstrate the impact de-novo protein design can have on disease intervention via therapeutics research and development.

Plants grow in designated areas known as meristems that contain stem cells. The gene LEAFY (LFY) is a flower meristem identity gene that initiates flower development in flowering plants and its homologs are also found in plants that do not produce flowers. Ferns are a lineage of land plants phylogenetically midway between the non-vascular bryophytes (such as mosses) and the seed plants. The model fern Ceratopteris richardii (Ceratopteris) contains two LFY homologs (CrLFY1/2). Given the described meristematic function of LFY in flowering plants, I am investigating whether CrLFY overexpression affects the development of the multicellular lateral meristem known as the “notch meristem”, the main site of cell division for the heart-shaped Ceratopteris gametophytes (the haploid stage of a plant’s life cycle). It has previously been established that CrLFY maintains cellular division in the apical cell of Ceratopteris gametophytes (a unicellular meristem that initiates development), but it is unknown if CrLFY also plays a role in notch meristem division once the apical cell terminates early in development. Consistent with our hypothesis, qualitative observations suggested that plants over-expressing CrLFY have larger and more U-shaped meristems, whereas wild type meristems appear more V-shaped. I am working to quantify the meristematic region within the notch in wild type compared to CrLFY over-expressing gametophytes using scanning electron microscopy and cell hand-tracing. I measure each traced cell and identify meristematic cells as those thinner and longer than the surrounding cells, with a length-to-width ratio of 3 or above. Additionally, I analyze the meristem angle to determine if there is a correlation between CrLFY expression and notch meristem shape. Understanding the role of LFY in plant meristems has important implications for the overall evolution of land plants, as master regulator genes such as LFY emerged in early plant lineages and evolved their functions over time.

A product we consume daily, such as honey, may contain heavy metals like copper (Cu). Copper can get into our environment and food from urban sources such as tires, roofing materials, and generators. We therefore expect food products from more urbanized areas to show a higher concentration of copper. To test this hypothesis we evaluated copper levels in raw honey from different regions with different levels of urbanization. The honeys we tested were divided into three different categories according to their source: rural, suburban, and urban. The samples were digested by reflux with HNO₃ and H₂O₂ then it was diluted and filtered in preparation for analysis by Atomic Absorption Spectroscopy. The results suggest that the honey with the highest concentration of Cu was from an urban area, and the honey from a rural setting had the least amount of Cu. A one-way ANOVA analysis confirmed a statistically significant difference in the copper levels in raw honey from the regions (p < 0.01). The concentration of Cu in the samples of honey we analyzed ranged from 0.2 ug/g to 0.4 ug/g which is well within the recommended upper limit of 300 ug/g. Since bees collect pollen and honey from plants within roughly one mile of their hive, evaluation of the levels of copper and other metals in honey can provide a snapshot of the background levels of exposure in that area.

Plants are generally unable to move reasonable distances as single adult organisms. This lack of mobility necessitates robust pathways that allow for response to environmental stimuli which can cause changes in the plant’s morphology to adapt to their changing environment. One such trait is flowering, and it is crucial for plant survival because it allows for plants to reproduce. The vast majority of agricultural plants are flowering plants, and robust growth and reproduction of these plants is especially important for the growing human population. This experiment aims to understand the interaction between an Arabidopsis thaliana nitrogen response gene named NITRATE-INDUCIBLE GARP-TYPE TRANSCRIPTIONAL REPRESSOR 1 (NIGT1), and an Arabidopsis gene which controls flowering called FLOWERING LOCUS T (FT). NIGT1 is known to modulate plant flowering despite being a nitrogen response gene. Here, I used a Dual-Luciferase Reporter System to test whether NIGT1 proteins directly interact with the FT promoter to regulate FT gene expression. In this system, the FT promoter is used to drive expression of Firefly Luciferase rather than FT. Separately, the NIGT1 gene coding region will be highly expressed under a constitutive 35S promoter. It will also be fused to the transactivation domain of the viral protein VP16 which converts transcriptional repression into transcriptional activation, as NIGT1 is known to be a transcriptional repressor. The VP16 transactivation domain will be repeated as four tandem repeats, forming the construct called VP64. The gene constructs of interest will be inserted into Nicotiana benthamiana using agrobacterium infiltration, then the leaf material from N. benthamiana will be used for the Luciferase assay. I hypothesize that the binding of NIGT-VP64 to the FT promoter will cause increased in Firefly Luciferase production compared to the absence of NIGT1-VP64. Higher amounts of Firefly Luciferase will result in greater luminescence, which we will quantify with a luminometer.

Plants recognize herbivore-associated molecular patterns (HAMPs) during herbivory that activates signaling to induce immune defenses. Caterpillar oral secretions contain Inceptin 11 (In11) which is a HAMP recognized by legumes such cowpea via Inceptin Receptor (INR). Thus, In11 and INR are a model system to study proteins involved in HAMP induced defenses, including Kunitz trypsin inhibitors (KTIs). It is known that KTIs are serine protease inhibitors with anti-herbivore activity; however, the precise role of In11 induced KTIs and the effect of cysteine content variation in cowpea KTIs remains unknown. Here, we show that selective removal of cysteines has a negative effect on KTI function in cowpea experiencing herbivory from the fall armyworm (Spodoptera frugiperda). We found that cowpea KTIs act as antiherbivore proteins against the fall armyworm when expressed in Nicotiana benthamiana, as we saw reduced weight gain on larvae feeding on leaves expressing wildtype KTI. Furthermore, we found that KTI function was negatively affected by the removal of cysteines, and larvae fed leaves expressing any of the mutant gained more weight than those feeding on wildtype. We hypothesize that these findings are due to reduced protein stability because we did not detect mutant KTIs in frass samples by westernblot. Understanding KTI protein structure and how it influences protein function is important for designing and selecting antiherbivore proteins to be used for plant defense in agriculture.

 The cGAS-STING pathway is a critical component of the innate immune system responsible for the detection of cytosolic DNA, which is key in identifying viral or bacterial infection. Activation results in the transcription of type I interferons (IFN), including IFN-β, and other proinflammatory cytokines that lead to the recruitment of immune cells to help control the infection. To explore this pathway, we use a system of conditionally immortalized macrophages (CIMs) derived from Cas9-transgenic i-TOMCAT mice. The Cas9 transgene allows us to make gene-specific knockout (KO) CIMs using CRISPR, while the i-TOMCAT allele, which drives expression of a fluorescent reporter (TdTomato) from the endogenous Ifnb1 locus, allows us to quantify type I interferon expression using flow cytometry. Using the STING agonist DMXAA, I found that CIMs in the progenitor state undergo rapid cell death upon STING activation, while terminally differentiated CIMs were resistant to cell death and produced high levels of IFN-β. Further experiments using the pan-caspase inhibitor Z-VAD-FMK and the necroptosis inhibitor necrostatin-1 showed that cell death in progenitor CIMs following STING activation is independent of apoptosis and necroptosis. This project aims to explore the signaling pathways downstream of STING that dictate either a cell death of IFN-β response. While considerable progress has been made in understanding how cGAS-STING signaling leads to IFN-β secretion, the cell death pathways associated with this cascade remain enigmatic. With recent studies showing that cGAS-STING activation can lead to tumor suppression by preventing cell division, promoting immunosurveillance, and triggering the release of tumor-specific antigens, research into this pathway may help guide the development of improved cancer immunotherapies.

Resilience in Alzheimer's Disease (AD) is defined by the difference between a person's expected and actual rate of cognitive decline given the severity of their disease. However, the mechanisms behind resilience are still unclear and I wanted to see if the anatomy of the brain over the course of AD could offer any clues. To accomplish this, cognitive tests and brain scans were obtained from patient data taken at Harborview Medical Center. Brain tissue atrophy in regions of interest were defined and combined into two measures. The first--biological subtypes--is whether the disease primarily affected the limbic regions or the cortical regions, while the second is left-right asymmetry. I found that resilience correlated with biological subtypes but not asymmetry. This suggests a way for us to predict resilience to better personalize treatment and eventually find ways to increase resilience.

The emergence and spread of antimicrobial resistance (AMR) in bacterial populations pose a significant threat to global health. A key factor in this process is the horizontal gene transfer (HGT) of resistance genes carried by mobile genetic elements (MGEs). This study focuses on understanding how mutations in plasmids influence the rate of HGT and its implications for the spread of AMR. Plasmids are small, extrachromosomal DNA elements that can transfer between different bacterial cells, a process crucial in spreading antibiotic resistance genes. These plasmids often use a mechanism known as conjugation for transfer, a process encoded by approximately 30 genes in the transfer (tra) operon. This research aims to understand how changes in the tra operon affect the plasmid's ability to move between bacteria. This movement can be either horizontal (between different cells) or vertical (from parent to offspring during cell division). Our project is divided into two main aims. The first is creating a library of mutated plasmids. I modified a large, well-studied conjugative plasmid, the F plasmid, and introduced specific mutations into the tra gene. This step is achieved using a method called Lambda Red recombination, a technique suitable for large plasmids where standard cloning methods fall short. The second aim is to measure the rates of vertical and horizontal gene transfer (VGT and HGT) in these mutated plasmids. We employed the Luria-Delbrück method for precise estimation of these transfer rates. Understanding the trade-off between VGT and HGT is crucial in predicting and controlling the spread of AMR. By mapping the relationship between specific mutations in the tra operon and their impact on gene transfer rates, our study contributes to a broader understanding of how antibiotic resistance spreads through bacterial populations. This knowledge is vital for developing strategies to combat the increasing threat of antibiotic-resistant bacteria.

Non-tuberculous mycobacteria (NTM) are ubiquitous in the environment and survive in various settings such as soil and water. The purpose of this project is to evaluate the ability of humoral responses produced from NTM pre-exposure on the prevention of Mycobacterium tuberculosis (M.tb) disease progression in mice. Additionally, we want to determine if antibody responses from immunization with bacille Calmette-Guerin (BCG) and/or candidate tuberculosis subunit vaccines contribute to protection against M.tb. To test this, we collected serum samples from mice that received various combinations of treatments before and after infection with M.tb. Groups either received NTM in the drinking water or normal drinking water, with or without BCG vaccine, and with or without a candidate vaccine or a combination of the treatments. M. avium ATCC 25291 NTM-lysate was used as an antigen for ELISAs and it was created by growing the mycobacteria and lysing the bacterial cells by multiple freeze-thaws. NTM-specific total IgG ELISAs were performed on serum samples following different vaccine strategies. Before M.tb infection, mice vaccinated with a candidate subunit vaccine regardless of NTM exposure or BCG vaccination had a significantly higher total IgG response. After M.tb infection, mice that received immunization with BCG followed by the candidate vaccine had significantly higher NTM lysate-specific total IgG responses compared to the saline group, however, this was observed only in NTM unexposed mice. NTM-exposed mice had equivalent levels of NTM-specific total IgG in the saline group when compared with the different vaccine regimens. This suggests that NTM exposure is able to produce potent NTM-lysate total IgG antibody responses in unimmunized ‘saline’ control mice, which suggests that the mice were likely boosted following M.tb infection. Given these findings, we plan to investigate whether the NTM-specific antibody response is reducing vaccine-mediated protective responses against M.tb.