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.

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.

Patients with Human Immunodeficiency Virus (HIV) are known to have increased risk of cardiovascular complications. High Density Lipoprotein (HDL) is a circulating lipoprotein responsible for removing lipids, such as cholesterol, from the blood and returning them to the liver, and is known to have a large impact on cardiovascular health. HDL is also known to have a protective effect on endothelial cells, which line the blood vessel walls, and it normally stimulates nitric oxide to cause an anti-inflammatory response. However, little is known about whether HDL from HIV patients has unique effects on the function of endothelial cells. I hypothesize that HIV-positive patients have increased inflammation due to impairment of HDL’s protective anti-inflammatory function. To test this, I am determining whether there is an increase in pro-inflammatory cytokines in plasma from HIV-positive patients compared to control patients, using enzyme-linked immunosorbent assays. I am also testing the hypothesis that HDL from HIV patients has a more pro-inflammatory effect on endothelial cells. I am culturing human microvascular endothelial cells (HMEC) and treating them with HDL from HIV-positive and non-HIV patients, along with appropriate control stimuli, followed by in-cell Western assays to measure activation of NFkB protein, a master pro-inflammatory regulator. I am using the same methods to measure activation of Akt, an intracellular signaling protein that can activate the production of nitric oxide via the enzymatic activity of endothelial nitric oxide synthase. I anticipate that HDL from HIV patients will cause increased activation of NFkB and decreased activation of Akt, which could explain, at least in part, the increased inflammation and cardiovascular issues in HIV patients. This research will begin to reveal possible mechanisms by which dysfunctional HDL may contribute to cardiovascular risks in HIV patients, and such findings could ultimately identify novel targets for therapeutic intervention.

Marine heterotrophic bacteria play a pivotal role in microbial community dynamics. This study aims to understand interactions within the microbial community of the oligotrophic (nutrient-poor) equatorial Pacific, specifically investigating how heterotrophic bacteria respond to the growth of autotrophic picophytoplankton. This experiment attempts to provide a more faithful representation of in-situ conditions, overcoming previous difficulties in capturing the dynamic behavior of microbial communities in the field. A novel methodology utilizing continuous chemostats with natural communities in the field accomplishes this objective. Three growth chambers were employed, two as chemostat systems and one in batch culture mode. All three growth chambers contained the natural microbial community that passed through a 3 µm pore-size filter. Chemostat systems continuously received 0.2 µm-filtered seawater (without microbes) while an equivalent volume was removed from the growth chamber simultaneously. Dissolved inorganic nutrients required by autotrophs – silicate, phosphate, and nitrate – were added to one of the chemostat’s 0.2 µm filtered media reservoir. This methodology was compared to a traditional batch culture, where nutrients were added to the growth chamber once, at time-zero. Cultures followed a 16-hour on/8-hour off light/dark cycle using LEDs, simulating the equatorial Pacific day/night cycle. Community responses were measured by continuous optical density measurements (OD), with endpoint subsamples analyzed for microbial abundance and DNA content using flow cytometry. Distinct day/night responses were observed in all cases, with the nutrient-enriched chemostat showing the most pronounced response. Overall, the results provide new insight into the linkages between marine autotrophic and heterotrophic microbes, while demonstrating an effective new methodology for examining microbial community responses to added nutrients. Thus, this study not only advances our understanding of microbial community dynamics in the oligotrophic equatorial Pacific but also introduces a novel experimental method that can be applied across a diversity of marine and aquatic environments.

Nutrients and CO₂ are important oceanographic variables, as they provide information which can be used to understand phytoplankton abundance and processes such as the oceanic carbon cycle. Therefore, as climate change impacts ocean systems, it is increasingly important to measure how nutrient and CO₂ concentrations in the ocean change over time and space. This study measured pCO₂ (which takes into account temperature, total CO₂, salinity, and alkalinity of the water), nitrate, phosphate and silicate concentrations in the western equatorial Pacific (5S-5N along 167W) in January 2024. Over space, pCO₂ and nutrients were analyzed for correlation with physical processes, primarily upwelling, using sea surface temperature (SST) and mixed layer depth. To determine the relationship of pCO₂ and nutrient concentrations to the biomass of microorganisms, correlations with fluorescence and beam transmission were also analyzed over space. Over time, pCO₂ was compared to atmospheric CO₂ and El Nino Southern Oscillation (ENSO) state to determine correlations between temporal pCO₂ trends and atmospheric phenomena. pCO₂ surface concentrations in the western equatorial pacific were found to have increased from 1983 to 2024 at an average rate of 2.02 +/- 0.034 ppm/yr and had a positive correlation with increasing average atmospheric CO₂ (R = 0.71, p-value < 0001). Spatially, surface pCO₂ and the macronutrients nitrate, phosphate, and silicate in the upper 200 m showed similar patterns from 5S to 5N along 167W. The concentrations of nitrate and phosphate had a significant negative correlation to mixed layer depth (R = -0.4, p-value < 0.001) and nutrients and pCO₂ had a significant negative correlation to sea surface temperature (p-value < 0.001). They peaked from 0-2N due to upwelling and exhibited smaller secondary peaks around 3S and 3N, likely due to mixing caused by north and south subsurface countercurrents. These results reinforce the importance of physical oceanic and atmospheric processes as a control for nutrient and inorganic carbon cycles in the western equatorial Pacific.

The nitrile imine produced by photolysis of 2,5 dimethyltetrazole undergoes a cross-linking reaction with the amide group in peptide-tetrazole conjugates and tetrapeptide-nucleotide complexes. In our work, we synthesized various peptide conjugates furnished with 2,5-diphenyltetrazole phototag. Upon laser pulses at 250 nm, nitrile imine intermediates can be generated by loss of N2 from tetrazoles. These nitrile imines can then crosslink with other parts of the molecule that contains amide groups. These crosslinking reactions are quite effective, achieving about 50% conversion with just two laser pulses at about 2 mJ. We could detect the formation of crosslinked products by tandem mass spectrometry. The UVPD-CID-MS3 spectra of these conjugates showed unique fragments including internal fragments of peptide sequence, indicating possible crosslinking. Moreover, we can confirm the structures and compositions of these crosslinked products using UV–Vis action spectroscopy and cyclic ion mobility mass spectrometry (c-IMS). By comparing experimental and calculated data, we confirmed the presence of nitrile imines and certain crosslinked products. We also explore thermal chemistry when nitrogen gas is lost from the peptide-tetrazole conjugates, and it seems to be a mildly energy-consuming process. The extra energy from breaking down tetrazoles is likely driving the reaction towards forming crosslinked structures involving peptide amide groups. Digging into the mechanism of this reaction, we found the proton transfer as the initial step, followed by a series of steps like cycloaddition and breaking of certain chemical bonds. Interestingly, other reactive groups, like cysteine thiol, do not interfere with this process. Within the complex of peptide conjugate and 2′-deoxycytidylguanosine, the intermolecular crosslinking efficiency is over 80%. The CID-MS3 and optimized structure showed the nitrile imine selectively targets guanine. In particular, the discovered reactivity of peptide amide groups toward nitrile imines appears promising as it provides potential clues to cross-link structure elucidation and conformational analysis.

Net primary productivity (NPP) is a major component of the carbon cycle. NPP is defined as the amount of carbon biomass produced by primary producers over a given period of time and area. The NPP exceeds 100 billion tons of carbon per year on Earth and half of it comes from the ocean through phytoplankton. The equatorial Pacific ocean is the largest tropical ocean on Earth and subsequently the largest oceanic source of CO2 to the atmosphere. Despite its importance, NPP in the west equatorial Pacific is poorly characterized due to the lack of data. Previous research suggests that strong upwelling is associated with increased nutrient concentration in the euphotic zone leading to an increase in primary productivity. However, the western equatorial Pacific is known for weaker upwellings compared to the eastern and central equatorial Pacific.This study was conducted aboard the R/V Thomas G. Thompson from December 28, 2023, to January 12, 2024, with the goal of identifying and quantifying the critical variables that have a substantial impact on NPP in the region including temperature, chlorophyll, dissolved nutrients, and current speed. NPP was measured using in-situ oxygen incubations, and was compared to NPP calculated from satellite data, which tend to typically overestimated or underestimated NPP in the region. Results of this study provide important information for refining satellite models to comprehend CO2 emissions into the atmosphere.

Each winter, the northeastern U.S. experiences powerful storms that cover cities in snow and ice, which result in millions of dollars in damage, halt travel, and disrupt essential services. Yet, the type, intensity, and distribution of precipitation is unique to each winter storm. This research project aims to provide a greater understanding of the precipitation properties and distribution in snowstorms, through focusing on a major winter storm that occurred over the Midwest on 17 February 2022 and was the target of a research flight conducted during the Investigation of Microphysics and Precipitation for Atlantic Coast-Threatening Snowstorms (IMPACTS) field campaign. Radar data collected during this research flight provides a unique perspective of the vertical cloud and precipitation structure, and numerical model fields provide the environmental context of the structures observed in the radar measurements. This storm had a frontal boundary, or a strong thermal contrast, that provided lift needed for the production of precipitation and had sub-freezing temperatures so that the precipitation fell as snow.This frontal boundary consisted of warm air originating from southern latitudes riding over colder air originating from northern latitudes. Analysis of the vertical cloud and precipitation structure from radar data and the in situ cloud particle measurements collected during the flight revealed that regions of higher reflectivity had larger particles and greater ice water content, compared to regions with lower reflectivity. The analysis also includes examining how the cloud particle properties are different depending on the origin of the air masses (from the north or south) that form the storm. By relating the temporal and spatial information regarding the air masses to the high-resolution radar and microphysics data collected by the IMPACTS airborne instruments, the results of this analysis will ultimately support increasing the accuracy of snow prediction.

In my research, I aim to analyze the impact certain tax global policies have on economic development depending on the countries' initial industrialization level. Using the development indicators published by the World Bank, among other sources, I will assess both a countries industrialization level and the impact of the tax policies. My research methods will combine theoretical economic frameworks with global, macroeconomic, and survey data. My unique approach will consist of statistical and historical data, looking into a variety of expert fields like health and education to assess the country. Thus, this paper will take an interdisciplinary approach to assessing economic conditions and policy impacts. Participating in this research will contribute to my transformative education by analyzing real world data and deriving policy recommendations. My work aims to advance our understanding of how tax policies impact both local and global development, with the ultimate goal of contributing to the design of more effective targeted tax policy. In this research, I expect one of two findings. Global tax policies that work well in industrialized nations work similarly or equally well in developing countries. Global tax policies that work well in industrialized nations fail to achieve the same results in developing nations. This concludes that tax policy recommendations have to be adjusted according to the industrialization level of the country.
 

Leigh syndrome (LS) is the most common form of mitochondrial disease in children. It affects 1 in every 40,000 births and its clinical manifestations include ataxia, seizures, failure to thrive and premature death. Genetic mutations in more than 75 different genes have been associated with LS. Among them is NDUFS4, the gene that codes for a subunit of the protein complex I of the mitochondria. Mice carrying a whole-body knockout (KO) of this gene greatly model this illness; they recapitulate multiple phenotypes of LS in patients. Prior studies in the lab have shown that the KO of Ndufs4 in GABAergic neurons, not in excitatory neurons, across all brain regions, reproduce the epilepsy phenotype seen in the global KO mice. Surprisingly, new KO mice with Ndufs4 inactivation restricted to GABAergic neurons of the brainstem and cerebellum interneurons, mediated by GlycineT2Cre, also have epilepsy. In this study, we sought to uncover the brain regions that house neurons involved in seizure activity in these mice. Brain regions experiencing neuronal hyperactivity during seizures in this new model of LS were examined. A thermal seizure was induced in the Ndufs4 GlycineT2Cre KO mice. For control condition, mice were exposed to a sham experiment. Forty-five minutes after the seizures or sham procedure, the mice were anaesthetized, and their brains were fixed and harvested. Brain slices were prepared and stained with a c-Fos antibody and finally imaged on the confocal microscope. Interestingly, high c-Fos immunoactivity was observed in the cerebellum alone and not in forebrain brain regions generally known to be involved in seizure generation. These findings indicate the participation of the cerebellum in seizure generation in Leigh syndrome epilepsy. In future studies we plan to increase the sample size and confirm the results with statistical methods.

Recent findings suggest that decreased chromatin acetylation leads to neurodegenerative diseases including Alzheimer's Disease (AD). Histone acetylation alters chromatin structure which regulates gene expression. One of the key proteins involved in this genetic modification is KAT5 (TIP60) acetylase. The Young Lab recently demonstrated that increased chromatin acetylation leads to the expression of genes involved in cognition and neuronal maturation. KAT5 is known to interact with the intracellular region of the Amyloid Precursor Protein (APP), which is a main player in the development of Alzheimer’s Disease (AD). In wild-type cells, the KAT5 protein is believed to be released from the membrane and translocated to the nucleus where it leads to increased chromatin acetylation and gene expression. Recent hypothesis suggest that an amyloidogenic cleavage of APP, either due to mutations or in AD conditions, prevents KAT5 nuclear translocation. To address this question, we will perform the following three experiments using human brain tissue and human induced pluripotent stem cell-derived neurons (hiPSC-Ns): 1. Immunohistochemistry for KAT5 on control and AD brains to see if we can detect a reduction in nuclear KAT5 localization in AD. 2. Immunocytochemistry on hiPSC-derived neurons harboring a familial AD (fAD) mutation in APP (Swedish mutation-K670N/M671L) as well as in neurons derived from cells that are deficient in APP (APP KO). 3. RNA-sequencing to determine differential gene expression between cells with fAD mutations and those that are deficient in APP, with a focus on genes regulated by KAT5. We expect to see aberrant KAT5 localization and gene expression in cells and tissues with AD and fAD mutations. Since treatments targeting the deposition of beta-amyloid led to many unsuccessful medical trials, we anticipate this study will demonstrate the importance of the absence of KAT5 signaling during early development of AD and devise new strategies for treatments.