Metabolic dysfunction–associated steatotic liver disease (MASLD) is a prevalent condition characterized by accumulation of fatty acids in hepatocytes, known as hepatic steatosis. While many factors contribute to lipogenesis and altered lipid transport, the molecular mechanisms driving lipid uptake remain under active investigation. One proposed mechanism involves regulatory RNAs that degrade transcripts of key lipid metabolism genes. Supporting this, lipid-accumulating hepatocytes express lower levels of AGO2, a core component of the miRNA-mediated mRNA regulatory machinery. To explore this, we developed a CRISPR-Cas9 knockout of AGO2 in human iPSC-derived hepatocytes. Following fatty acid challenge, the lipogenic and lipid-binding genes MLXIPL, SOAT2, and FABP5 were upregulated, while the lipid-transport gene TM6SF2 was downregulated, suggesting that miRNA-mediated regulation of these genes helps maintain cellular lipid homeostasis. This project aims to identify the specific miRNAs targeting these genes, with the long-term goal of uncovering therapeutic targets to restore healthy lipid metabolism. miRNA profiling of the AGO2-knockout line revealed downregulation of miR-432, miR-136, and miR-127—suggesting they may normally suppress the genes upregulated in lipid-accumulating cells. Conversely, miR-16, miR-26, and miR-184 were upregulated, consistent with an opposing role on lipid metabolism gene expression. Further analysis is required to confirm their involvement in the lipid imbalance phenotype.

X-linked agammaglobulinemia (XLA) is a primary immunodeficiency caused by loss-of-function mutations in the Bruton Tyrosine Kinase (BTK) gene, resulting in impaired B-cell development and absence of circulating immunoglobulins. Current treatment with immunoglobulin replacement therapy improves survival but requires lifelong administration and does not fully restore immune function. This project investigates whether precise genome editing can enable durable and physiologically regulated correction of BTK in hematopoietic stem and progenitor cells (HSPCs). We developed an adenine base editing approach to correct the pathogenic BTK c.763C>T mutation without double-stranded DNA breaks. Guide RNAs were screened with PAM-flexible ABE8.20 variants in a K562 model and primary human CD34+ HSPCs to identify optimal conditions. ABE8.20 paired with gRNA4 achieved up to 88% A-to-G conversion in primary HSPCs, demonstrating efficient correction of BTK. To address delivery limitations and move toward in vivo genome editing, we evaluated mini enveloped delivery vehicles (miniEDVs) as a non-integrating alternative to electroporation. VSV-G pseudotyped miniEDVs achieved up to 50% genome modification in primary CD34+ cells using Cas9 RNP delivery, while miniEDVs packaging adenine base editors showed dose-dependent editing in vitro, reaching ~6% in K562 cells. Together, these findings support the development of a combined base editing and delivery strategy toward durable, non-integrating, and potentially in vivo gene correction for XLA.

Trypanosoma brucei is a severe human and animal pathogen that remains a danger in large areas of Africa. It is a heteroxenous, parasitic protozoan, with mammals and the tsetse fly as its primary reservoir and vector respectively. The parasite requires mobility to infect and complete its life cycle, so the flagellum and its proteins, vital for movement, have been studied extensively. We have researched novel microtubule inner proteins (MIPs) and microtubule outer proteins (MOPs), have unknown effect on T. brucei mobility. The project focuses on MC1, a MOP that lines the microtubule. First, the protein was tagged with neon green GFP in vitro in the 2913 wild-type T. brucei brucei in their procyclic fly form by integrating a plasmid into the cells through electroporation. Cells that took up the DNA were selected for by a drug marker (puromycin). After that, a knockdown plasmid was integrated into the ngGFP-tagged 2913 strain tryps. This uses a RNAi silencing system. The knockdowns were selected for with a drug (Blasticidin). The tryps were tested for presence of MC1 protein through Western Blot. Later, motility assays will be carried out to detect a defect in motility in the mutants. A growth curve will also be done to detect a division defect. Data analysis is still being done, so results are pending.

Alphavirus is a genus of (+) sense single-stranded RNA viruses that can cause multiple arthritic and encephalitic conditions in vertebrates including humans. Alphaviruses pose a significant, growing risk to global human health, and investigating the host immune responses to them would provide essential information to curbing further spread. At the molecular level, cells can rapidly discern virus-specific signatures, activate immune signaling pathways, and suppress viral replication. One such mechanism relies on the host zinc finger antiviral protein (ZAP), an RNA-binding protein and a potent restrictor of alphavirus translation. ZAP recruits an E3 ubiquitin ligase called tripartite motif protein 25 (TRIM25) to enable its antiviral function. TRIM25, once recruited, facilitates the ubiquitination of multiple proteins, a process with various biological consequences including targeting substrates for degradation, activation, relocalization, signaling, and more. How TRIM25-mediated ubiquitination contributes to the ZAP antiviral response is not well understood. Preliminary results from mass spectroscopy proteomics suggest that TRIM25 ubiquitinates all four viral nonstructural proteins (nsP1-4) of Sindbis virus, a model alphavirus, during infection. We subsequently used a targeted overexpression system to confirm that TRIM25 interacts with nsP3. This project aims to further establish nsP3 (and other nsPs) as a bona fide substrate for TRIM25, and determine the functional consequences of nsP ubiquitination on viral replication.

Monkeypox virus (Mpox) is a double-stranded DNA virus that results in a rash and flu-like symptoms. While its fatality rates are relatively low (3-10% for clade I and less than 1% for clade II), continual monitoring for increased pathogenicity and mortality remains of utmost importance. Wastewater genomic surveillance is a powerful tool for the identification and monitoring of pathogens circulating in the community. My project aims to detect pan-clade Mpox in longitudinal wastewater samples from Los Angeles County using quantitative polymerase chain reaction (qPCR). First, I used bead-based methods to enrich for viral pathogens and isolate DNA and RNA from weekly wastewater samples collected from October 6, 2025 to February 16, 2026. Then, I synthesized cDNA from the extracted RNA. Finally, I will perform qPCR to amplify and quantify Mpox and pepper mild mottle virus, a normalizer for fecal pollution. By comparing the Cq values yielded by qPCR of the weekly wastewater samples to those of known standards, changes in Mpox abundance over time can be assessed. Total DNA, RNA and cDNA quantities have shown similar fluctuations over time, suggesting changes in the general abundance of circulating pathogens as well as successful cDNA synthesis from RNA. I hypothesize that Mpox will be detectable in our wastewater samples from November 3 to December 15, 2025, with quantifiable changes proportional to reported hospitalizations. This work can provide insight into the ways in which Mpox fluctuates as outbreaks emerge and subside.