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Sona Kang , Department of Nutritional Sciences and Toxicology 

"Examining ten-eleven translocation methylcytosine dioxygenase 1-directed regulation of thermogenic marker gene expression" 

Abstract: 

"Obesity, characterized by the excessive accumulation of body fat, and associated metabolic disorders, like type 2 diabetes mellitus, present major health concerns that affect a large portion of the global population. Adipose tissue, also known as body fat, is divided into two known categories of mammalian adipose tissues––white adipose tissue and brown adipose tissue (BAT). BAT plays a large role in thermogenesis, the process by which energy is dissipated as heat, and studies have suggested that impaired thermogenesis may relate to obesity. Generally, brown adipocytes highly express UCP1, which is a mitochondrial membrane protein with key functions in thermogenesis. Subcutaneous white adipocytes can undergo a “browning” process to become beige adipocytes and obtain thermogenic functions. Given this potential to activate thermogenic function, beige adipocytes are being researched as a therapeutic target for obesity and other metabolic disorders. A prominent facet of the browning process is the upregulation of thermogenic genes, which implicates shifts in chromatin structure and involvement of key regulator proteins in gene expression. At the Kang Lab, we are interested in the ten-eleven translocation methylcytosine (TET) dioxygenase protein family, which is comprised of proteins that can catalyze demethylation of 5-methylcytosine and interact with gene expression through transcriptional regulation. Identifying key players in epigenetic control of thermogenesis has important implications for the future of therapeutic approaches. This particular study elucidates the effect of Tet1 on the expression levels of thermogenic marker genes, including Ucp1, Ppargc1a, etc. Through a loss-of-function model and associated experiments in beige adipocytes, we have suggested that Tet1 plays a repressive role in thermogenic marker gene expression. Additionally, we have demonstrated that Tet1 knockdown significantly increases mitochondrial respiration, thereby implicating Tet1 as an important target for future therapeutic approaches for obesity and other metabolic disorders based on an overexpression paradigm that increases energy expenditure."

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Rong Wang , University of California, San Francisco 

"Alk1 deficiency initiated arteriovenous malformations by enlarging capillaries and arteries" 

Abstract: 

"Vascular perfusion from arteries to veins through capillaries is important throughout the body, but this is impaired in patients with arteriovenous malformations (AVMs). AVMs are vascular lesions that shunt blood from arteries to veins and can lead to ischemia and hemorrhage. It is the primary manifestation of Hereditary Hemorrhagic Telangiectasia (HHT), known to be caused by Alk1 mutations, and its formation processes are not well understood. Our lab has developed a novel mouse model of Alk1-mediated AVMs. In this model, we show that AVMs form through the initial enlargement of capillary and arterial vessels using both time-lapse two-photon imaging, as well as a newly developed sensitive image quantification approach. Furthermore, our evidence does not show that increased endothelial cell (EC) proliferation in these vessel segments drives the increase in vessel diameter. Understanding the primary changes that occur during AVM formation will provide insight into the causative events and role Alk1 in disease development and may serve as therapeutic targets. "

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Michael B. Eisen , Department of Molecular and Cell Biology, University of California, Berkeley 

"Examining Internal Representations of Protein Disorder in Convolutional Neural Networks" 

Abstract: 

"Disordered regions within proteins adopt a variety of structures and have amino acid sequences that can vary dramatically between homologous proteins in different species. Currently, the relationship between sequence and function in disordered regions is poorly understood as the lack of primary structure conservation prevents the identification of homologous disordered regions and the transfer of functional annotations between them. However, recent studies have reported evidence of “molecular features” which are conserved in disordered regions. Notably, patterns of these conserved features are associated with specific functions permitting the prediction of function from sequence alone. Unfortunately, because these features were curated from previous studies, many have overlapping definitions or measure similar quantities. Thus, they likely do not represent a complete catalog of all disorder-associated features. Given their success at predicting the functional properties of proteins, our lab hypothesized that a neural network trained to identify disordered regions would learn efficient and complete representations of disorder-associated features. Using a data set I compiled from publicly available structural study data, I trained several convolutional neural networks (CNNs) which predict disorder at the level of individual residues. The features learned by these CNNs were extracted and compared to known disorder-associated features calculated from the same sequences. This analysis showed that many learned features were local in nature and correlated with certain known features. In particular, strong associations were found between learned features and known aliphatic, hydrophobic, aromatic, and proline-associated features highlighting their relevance for describing disorder and demonstrating the ability of CNNs to carry out effective feature selection. Unique learned features strongly correlated with known serine-associated features were also observed, indicating that CNNs likely also learn different types of disorder. These results demonstrate that CNNs do encode meaningful representations of disordered regions and validate the use of this method to identify novel disorder-associated features.  "

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Daniel K Nomura  , Innovative Genomics Institute, University of California, Berkeley 

"Rational Design of Molecular Glue Degraders" 

Abstract: 

"Since the mid-20th century, the field of drug discovery for disease regulation has focused on the concept of occupancy-driven pharmacology, a process in which a protein’s function is inhibited due to an organic small molecule binding at the active site of a protein. However, over 90% of the proteome is currently deemed “undruggable” due to a lack of surface-lying ligandable hotspots. Many of these undruggable proteins, such as transcription factors and protein complexes, are implicated in disease but cannot be targeted through active site directed inhibitors, creating a need for new therapeutic modalities that can control protein function. One method for targeting undruggable proteins is through the use of molecular glue degraders, small molecules that chemically elicit a novel protein-protein interaction between a component of the ubiquitin protease system and a neo substrate protein, resulting in the degradation of the protein target. Using the CDK4/6 inhibitor ribociclib, the Nomura Research Group synthesized various analogs of the drug bearing appendages at its secondary amine exit vector to develop a novel molecular glue. Through immunoblotting and chemoproteomics, it was confirmed that molecules EST1027 and EST1060 brought target protein CDK4 in close proximity to E3 ligase RNF126, causing CDK4 protein degradation. I sought to determine the essential chemical elements that define molecular glue degraders, and use these findings to develop a chemical moiety that can be appended onto a protein inhibitor to confer molecular glue degrader properties to it. Throughout my project, I determined the minimum recognition elements of EST1027 and EST1060 that was required to bind to RNF126, and append this moiety onto protein targeting ligands to cause degradation of other therapeutically relevant protein targets. My project will make it easier to develop molecular glues for disease-related proteins currently deemed undruggable by current pharmaceutical techniques. "

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Britt Glaunsinger , Department of Molecular & Cell Biology 

"Elucidating the role of ORF6 transcriptional activity in Lytic Replication of Kaposi's sarcoma-associated herpesvirus (KSHV)" 

Abstract: 

"Kaposi sarcoma-associated herpesvirus (KSHV) is a double-stranded DNA oncogenic virus that disproportionately affects immunocompromised individuals. My thesis work focuses on ORF6, an essential KSHV protein that past researchers have shown plays two roles in the KSHV life cycle: transcriptional activation and replication of the viral genome. My project is on the relationship between these two functions and evaluating if ORF6 transcriptional activity contributes to viral DNA replication. To test this hypothesis, I used CRISPR Cas9 to truncate ORF6 and remove its activation domain and tested if KSHV lytic viral DNA replication is reduced. In collaboration with my mentor, I observed reduced levels of vDNA replication, supporting the hypothesis that ORF6 transcriptional activity contributes to viral DNA replication. However, when I conducted an ORF6 western blot to detect the truncated form of ORF6, I instead observed depletion of ORF6 protein. This indicates that reduced levels of vDNA replication may be from ORF6 protein depletion rather than the ORF6 activation domain being truncated. Thus, I evaluated the use of Cas9 mediated Homology Directed Repair (HDR) to allow me to install precise mutations in key residues of the ORF6 activation domain rather than removing the entire domain. To measure and optimize the ability to perform HDR, I am conducting a pilot study that replaces the GFP cassette on the viral genome with the blue fluorescent protein. Initial experiments indicate that while GFP is efficiently targeted, no HDR is occurring as cells did not fluoresce blue. This may be from low efficiency delivery of the HDR template using transfection reagents, and I am currently exploring alternatives like nucleofection for template delivery. By allowing precise mutations of ORF6 that mutate the activation domain while preserving the ORF6 protein, I can better evaluate the relationship between the transcriptional activation and viral lytic replication function of ORF6."

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Bethanie Edwards  , Department of Earth & Planetary Sciences 

"Lipids as bioindicators of nutrient availability and temperature in a changing climate" 

Abstract: 

"Ocean “omics” aims to characterize the ocean by genetic and molecular profiles, including DNA, proteins, and metabolites. Application of these strategies is an important tool for marine conservation, allowing researchers to distinguish and monitor marine life. Lipids comprise biological membranes, their structure influenced by nutrient availability and temperature comprising significant criteria while monitoring climate change in marine ecosystems. We performed a trophic study in the Pacific Ocean, collecting samples of phytoplankton and zooplankton at 19 stations in the Pacific Ocean representing 19 latitudinal zones. Each latitudinal zone is placed into one of three geographic regions: the Hawaii Coast (HC), the North Pacific Subtropical Gyre (NPSG), and the California Coast (CC). We analyzed the 19 phytoplankton and zooplankton lipid samples respectively using high-performance liquid chromatography coupled with mass spectroscopy. We extracted DNA from all zooplankton samples for species level identification of each geographic region. We focused on fatty acid saturation varying with temperature and phospho-free lipid composition varying with geographic region upon statistical analysis. We found that fatty acid saturation is intrinsically linked with temperature, and the presence of phospho-free lipids increases in the NPSG. Nutrient availability is affected by altered wind and water circulation, increasing or decreasing the availability of essential nutrients to marine life. Temperature fluctuation has broad implications to the health of marine ecosystems, contributing to sea-level rise, coral bleaching, and loss of marine biodiversity. Lipids serve as important bioindicators to the stability of marine ecosystems, especially pertaining to nutrient cycling and temperature fluctuation."

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Timothy Henrich   , Division of Experimental Medicine, University of California, San Francisco 

"Investigation of CCR5 Inhibition on the Immune Response and HIV Persistence Post-Transplant" 

Abstract: 

In an effort to understand the dynamics of solid organ transplantation for people living with HIV, there is an opportunity to explore the immune responses and HIV-1 reservoir of patients after a transplant procedure. Several studies within the past decade have reported that patients with HIV-1 have higher rates of graft rejection post-transplant compared to their counterparts, yet the reasoning for this phenomenon remains unclear. Furthermore, the HIV-1 reservoir post-transplant is not well characterized, and the mechanisms that enhance HIV-1 persistence in CD4+ T cells despite highly active antiretroviral therapy (HAART) are not fully understood. Clinical studies have provided evidence that suggest chronic inflammation and T cell activation aid in HIV-1 persistence in CD4+ T cells. In a recent 2014 transplant study, the C-C chemokine receptor type 5 (CCR5) was targeted in order to determine the impact of modulated immune regulation on proviral DNA levels in CD4+ T cells. Evidence gathered suggests that reduced CCR5 expression is associated with decreased levels of proviral HIV-1 DNA. In this ongoing study, we are examining a cohort of 150 participants with HIV-1 following kidney transplantation. Maraviroc, a CCR5 antagonist, will be used as an immunosuppressant in combination with ART to determine the impact of CCR5 inhibition on the immune response and HIV-1 persistence. Analysis of the immune system will begin by first evaluating protein expression at timepoints 1 and 6 months after transplant. HIV-1 persistence will be assessed by analyzing peripheral blood mononuclear cells (PBMCs) sampled at 1, 3, 6 and 12 months post-transplant. The intact proviral DNA assay (IPDA) as well as RT-PCR will be conducted to measure replication-competent HIV-1 proviruses and transcriptional activity over time in CD4+ T cells, respectively. The data provided through this study has the potential to direct strategies for post-transplant outcomes, as well as further research on elimination of HIV-1 in reservoirs.