Prostate cancer has a significant heritable component. It is estimated that 10-15% of patients with advanced prostate cancer carry an inherited predisposing genetic mutation, and these patients generally present with a younger age of onset and a strong family history of cancer. The standard in the field of oncology is to conduct short-read DNA sequencing on such patients to find predisposing mutations. While short-read sequencing does well to identify simple mutations that cause disease in many families, our lab concluded that short-read sequencing misses critical mutations in many prostate cancer susceptibility genes. We hypothesized that prostate cancer in many families is due to complex inherited mutations such as genomic deletions, inversions, and mobile element insertions that are not detectable by conventional genomic technologies such as short-read sequencing. To test this hypothesis, our lab specifically recruited prostate cancer patients who, despite having family histories of cancer, did not have any mutations detected via conventional genetic sequencing methods. This project utilizes Nanopore long-read DNA sequencing, which reads DNA in longer fragments and can reliably detect complex mutations. My role is to conduct long-read sequencing on DNA samples from these patients, then analyze the DNA sequence for mutations. I have sequenced 32 patients so far and identified 4 complex mutations through long-read sequencing which were missed by other approaches. These complex mutations include insertions of repeat sequences and duplications which disrupt gene function in BRCA1 and BRCA2. This suggests that, consistent with our hypothesis, some patients who do not have mutations found via conventional sequencing methods do indeed carry causative mutations in well-established prostate cancer risk genes. By finding these mutations, patients can receive more targeted and effective cancer treatment, and undiagnosed family members stand a better chance of catching cancers at earlier stages.

In this experiment we will look into the relationship between different mushroom species' toxicity and copper concentrations. Due to their wide variety of biochemical characteristics, mushrooms can be either extremely toxic or edible. Mushrooms contain different amounts of copper, an essential trace element that may affect a mushroom's toxicity. Using Atomic Absorption Spectroscopy, we evaluated the amount of copper of several mushrooms. Our early results show that mushrooms with higher copper concentration tend to be more toxic. This shows that copper content may be a useful marker of the toxicity of mushrooms, giving foragers important information and assisting in the development of food safety protocols. Our research is to be continued as we’re going to test on more mushrooms to get a better understanding on how copper could affect the production of toxic compounds.

Recent studies have reported that certain tampon brands contain traces of various metals, raising public safety concerns about regular tampon use. Exposure to metals such as lead may pose detrimental effects on cognitive function, the nervous system, and reproductive health, yet little is known about the extent to which these metals are absorbed into the bloodstream from these products.  This project aims to investigate the presence of heavy metals within tampons. We hypothesized that tampons made from cotton would contain higher traces of metals compared to ones that are made with viscose rayon. We selected five widely available brands of varying absorbances and material, categorizing them as either organic (cotton) or non-organic (viscose rayon). To quantify the total lead content, 0.300 g of each sample was digested using a mixture of hydrogen peroxide and nitric acid. To determine the extractable quantity of lead, each sample was submerged in a simulant solution for 24 hours, replicating the acidity of vaginal fluids. To ensure the presence of lead within the sample, tampons with measurable lead concentration were spiked with known amount of lead quantity. Using AA Spectroscopy, quantifiable total lead contents were found in three out of the five tampon samples; Tampon C exhibited the highest lead content of 1.363 µg/g of tampon. Additionally, only one in five tampon samples was found to have significant extractable lead content, with Tampon C containing 0.2184 µg/g of tampon. Our results indicate a higher proportion of detectable traces of total and extractable lead in non-organic tampons compared to organic tampons. Despite these findings, further research is needed to establish whether there are adverse health effects to lead exposure from tampon use.

Methane is one of the most attractive targets for controlling near-term climate change due to its short lifespan and high potency (34 times that of CO₂). Methanotrophs are bacteria that can consume methane and convert it into CO₂ and biomass. There is growing interest in using these bacteria to mitigate greenhouse gas emissions from sources such as landfills, agricultural feedlots, and abandoned coal mines. However, a key challenge is that to achieve large scale methane sequestration, as well as economic viability of deploying these in the field, we have to significantly improve the growth of methanotrophs at low concentrations of methane. Regulatory genes play an important role in determining how bacteria allocate energy. By deleting specific regulatory genes and measuring the growth rate of these mutants under low methane conditions, we can assess their importance in helping the bacteria survive and thrive in nutrient-limited environments. Using this approach, we can also replicate mutations that have naturally emerged in strains cultivated for over a year under low methane conditions. This allows us to confirm whether these mutations provide a growth advantage. By identifying and testing key genes involved in low-methane growth, we are guiding efforts to engineer a more efficient and resilient strain for real-world applications.

Neuropsychiatric disorders lead to devastating impacts on a patient’s life, affecting physical movement, cognition, and behavior. A pattern observed in patients with neurodegenerative disease includes neurofibrillary tangles in the brain, which may be caused by the abnormal accumulation of the microtubule-associated protein tau (MAPT). Tau is encoded for in the MAPT gene locus on chromosome 17, where the locus commonly interacts with an enhancer to boost transcription. However, some patients have one copy of chromosome 17 with an inversion that breaks this interaction, which is associated with lower risk of disease. This raises the question of whether the MAPT locus interacts with other enhancers that increases tau production. Thus, the goal of this project is to identify genetic variants that influence the 3D interactions between the MAPT locus and potential enhancers in patients heterozygous for the MAPT inversion associated with a lower risk of neuropsychiatric disorders. To accomplish this objective, neuronal nuclei extracted from patients are analyzed using fluorescent in-situ hybridization (FISH) to identify interactions in the MAPT gene locus. We will map a 2 Mb region of chromosome 17 centered on the inversion using FISH probes. This region is broken up into ten 200 kb spots to be individually visualized using fluorescent oligonucleotides through a fluidics system, to create a composite image of all spots. Interactions involving the MAPT locus may be identified by comparing distances between spots, in which gene segments that interact would have a shorter distance compared to segments that do not interact. This would allow us to find genetic variants associated with the chromosome 17 inversion that potentially influence MAPT gene regulation.

The Lidstrom Lab aims to better understand methane-consuming microbes (also called methanotrophs) so that we can develop technologies to remove anthropogenic methane emissions, which will reduce the severity of global warming. Our research explores how the methanotroph Methylotuvimicrobium buryatense 5GB1C can be bioengineered to grow well at the low methane concentrations found in human-made emission sites, while providing value-added products like biomass from dead bacteria that can be used as animal feed. Understanding bacterial methane utilization will allow us to create effective biocatalysts at a far lower monetary and environmental cost. My research project involves deleting cytochrome genes that may be important for the 5GB1C strain to grow in low methane conditions. Manipulating these genes may allow for further improvement of growth at low methane. My targets are three genes that encode cytochromes, which are electron carriers that take electrons from particular reactions and supply them to other reactions that are otherwise energetically unfavorable. My hypothesis is that these cytochromes are involved directly in supplying 5GB1C with electrons needed for the oxidation of methane into methanol. If these cytochromes supply electrons required for methane consumption at low methane, then deleting them would generate a mutant that would grow poorly on methane because it lacks the electron carrier(s). I have generated two possible cytochrome deletion mutants and continue to work on a third cytochrome. Once the mutants that can be generated are sequenced to verify the deletions, cultures will be grown under low methane and methanol conditions to determine how their ability to grow has been affected by the knockout mutations. In this manner, our lab is building a valuable knowledgebase of genes that are suitable for manipulation to improve growth in low methane for the technologies that one day will help curtail the worsening of global warming.

Methane is an extremely potent greenhouse gas, with a warming potential 86 times greater than that of CO2 on a 20-year timescale, and is therefore a top priority for mitigation efforts to combat climate change. Methanotrophic bacteria, such as M. buryatense 5GB1C, metabolize methane as their main source of carbon and chemical energy, a trait that could help slow climate change by reducing emissions. A major obstacle is the rate at which methane consumption occurs at low methane concentrations, which tends to be too low to be appreciable. This project seeks to answer whether currently unknown genes involved in the growth of M. buryatense 5GB1C on low methane could be discovered by comparing its genome with that of a closely related methanotroph, M. alcaliphilum 20Z. While the two have very similar genomes and metabolisms, M. alcaliphilum is not able to grow at low methane concentrations (500 parts per million), while M. buryatense is. I analyzed the two genomes and isolated all genes present in M. buryatense without homologs in M. alcaliphilum. Because they are unique to M. buryatense, they may be involved in the observed growth difference. I systematically performed targeted deletion mutations on many of these candidate genes, and then tested them for growth on low methane compared to the wild type strain, looking for any defect that would suggest a gene directly essential to growth at 500ppm. I confirmed several genes to have no impact on growth at low methane, as well as one that appears to be essential to growth in any conditions, and anticipate reaching conclusions on several more mutants. These findings will help to develop microbial methane mitigation technologies that can be utilized in a great range situations and at a larger scale, essential characteristics for a global impact.

Protein Kinase Inhibitors (PKIs) are a family of heat stable, high-affinity inhibitors of the catalytic subunit of Protein Kinase A (PKAc). In the presence of Mg-ATP, the three isoforms—PKIα, PKIβ, and PKIγ—bind to PKAc with very low dissociation constants: 0.758nm, 1.875nm, and 0.4142nm respectively. In vitro studies have shown that PKIs can translocate PKAc from the nucleus to the cytoplasm, suggesting a role for PKIs in terminating nuclear cAMP-driven PKA activity. Previous research, including studies from our lab, has found that dysregulated PKAc mutants play a significant role in Cushing’s syndrome, a rare and potentially fatal metabolic disorder caused by excessive cortisol production. Building on these findings, we hypothesized that increasing PKI expression could counteract the hyperactivity of PKAc mutants and reduce cortisol production. To test this, we expressed each PKI isoform in adrenal cell lines and assessed their steroidogenic capacity using biochemical assays such as western blots, RNA-seq, qPCR, and ELISA-based cortisol assays. We observed that PKIα and PKIγ led to a general suppression of steroidogenic associated proteins such as StAR, Cyp11a1 and SF1. This altered proteome was accompanied by significantly suppressed cortisol synthesis only in the PKIα and PKIγ expressing cells. The difference between PKIα/γ and PKIβ was surprising given that all PKI isoforms are postulated to potently inhibit PKAc. Thus, we questioned whether PKIα/γ effects are mediated through PKAc. To answer this, we have cloned mutant PKI isoforms that do not bind PKAc, and confirmed the mutant PKIs do not inhibit PKAc through kinase assays. Our next step is to express the mutant PKI isoforms in adrenal cells and assess their effect on steroidogenic capacity of the cells. Our findings suggest that PKIα and PKIγ play key roles in cortisol regulation and may have broader implications for gene regulation in adrenal cells.

Young adult cannabis use has become increasingly prevalent in the US, particularly among individuals attending four-year colleges. The perceived social acceptability of cannabis use plays a crucial role in shaping attitudes and behaviors towards substance consumption. While societal attitudes towards cannabis have evolved over the last two decades, there is a gap in understanding how these perceptions differ between college students and their non-college peers. My research aims to compare perceptions about the social acceptability of cannabis with the actual frequency of use among young adults who attend four-year colleges, versus same aged individuals that are not attending school. I am using a subsample of young adults using baseline data from a larger longitudinal study on health behaviors, the Washington Young Adult Health Survey (WYAHS), for the analysis. I am conducting the data preparation and analysis using SPSS. I believe that there will be a significant difference in perceived social acceptability of cannabis use between college students and those not attending school, but I also anticipate that actual consumption will not be significantly different. The results of this research could be important for improving substance use education and addressing preconceived notions of cannabis use acceptability among young adults. Previous research on the WYAHS data has shown significant changes in substance use behaviors over the last six years, especially throughout the pandemic. Future research is needed, which focuses on how my findings may change when based on data from before the COVID-19 pandemic.