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2019 Research Showcase
MR Abstracts
Advisor : ETHAN BIER
Abstract Title : A Mechanistic Understanding of the Black Elusive Eschars in Cutaneous Anthrax
Abstract : The severe and often lethal Anthrax disease has been identified since antiquity, and is caused by the spore-forming, gram-positive bacterium, Bacillus anthracis (B. a). Anthrax in Greek translates to coal, coined from the black skin lesions conventionally seen by those with cutaneous anthrax. The cause of these black eschars is not understood nor has been adequately investigated. B.a secretes Edema toxin (ET), which paralyze the immune response in the early phase of infection and produce aberrant symptoms in later stages of the disease. ET is a potent Adenylate Cyclase, that causes the uncontrolled rise in cAMP concentrations, and is responsible for edema, and other symptoms. Previous literature in melanocytes has shown cAMP signaling facilitates pigmentation through a defined pathway that results in melanin trafficked to exterior keratinocytes. Moreover, activation of Protein Kinase A (PKA) by cAMP, leads to specific phosphorylation and activation of Tyrosine Hydroxylase (TH) the rate-limiting enzyme required for melanin and dopamine synthesis. Exploitation of TH in Drosophila melanogaster established a direct relationship with pigmentation, allowing us to further hypothesize EF’s direct effect on pigmentation through TH. A mammalian-cell model system further supported the direct TH relationship with pigmentation.
Advisor : BRYAN K. SUN
Abstract Title : HOPX is a ZNF750 Target that Promotes Late Epidermal Differentiation
Abstract : The outermost layers of skin and oral mucosa consist of stratified epithelia that serve as a barrier and interface to the outside environment. These epithelia are comprised primarily of keratinocytes, which exist as self-renewing progenitors in the innermost layer that differentiate into overlying protective layers. On a transcriptional level, keratinocyte differentiation involves coordinated induction and repression of hundreds of genes, but the genetic regulators governing this process are not fully understood. The homeodomain-only protein homeobox (HOPX) is known to be involved in keratinocyte differentiation, but prior studies disagree whether it promotes or inhibits this process. Here, we show that HOPX activates skin differentiation in primary human keratinocytes and organotypic epidermal tissue. HOPX expression is induced by the transcription factor ZNF750, which binds an enhancer located ~6 kb downstream of the 3’ end of the HOPX gene. Whole transcriptome analysis demonstrates that HOPX controls expression of 561 differentially expressed genes and is a positive regulator of late epidermal differentiation. These results show that HOPX functions downstream of ZNF750, a well-defined transcription factor in the skin that is activated by p63. In this context, we propose that HOPX functions within a p63-ZNF750-HOPX pathway to upregulate key proteins required for terminal epidermal differentiation, providing clarity to previous studies showing conflicting results. Future studies will aim to decipher the molecular pathways and mechanisms operating downstream of HOPX that engage the late differentiation program.
Abstract Title : Ectotherms and the Temperature Size Rule
Abstract : Dubbed Bergmann’s size cline, animals will naturally develop larger body sizes at higher latitudes and thus lower temperatures, and smaller body sizes at lower latitudes, and thus higher temperatures. This makes sense for endotherms, however ectotherms seem to be much more varied in their responses. In a laboratory setting this phenomenon is know as the Temperature Size Rule (TSR). Within laboratory confines, this must be due to a phenotypically plastic response. Many hypotheses have been proposed as to why ectotherms respond to temperature as they do, and a central question revolves around whether or not these response are adaptive or nonadaptive. One mechanistic proposal by Kammenga et al. illustrates that a single nucleotide polymorphism in tra-3, a calcium dependent protease, shows strong correlations with temperature responses in C. elegans.
Abstract Title : Comparing dendritic morphology and axon projections of excitatory and inhibitory neurons in the Xenopus laevis optic tectum
Abstract : The brain contains a diverse network of neurons that vary in complexity. Investigating the properties and connectivity of these neurons is essential to identifying the relationships between brain function, behavior, and neurological disease. A useful animal model for studying the brain is the albino Xenopus laevis tadpoles due to their transparent skin and visible external development. One region of interest is the optic tectum, a midbrain region homologous to the mammalian superior colliculus, which is responsible for visual processing and avoidance behavior. Previous studies have shown the optic tectum to be responsible for these behaviors, but knowledge is still limited on what neuronal cell types exist in this region and how these neurons interact with each other to operate behavior. In order to characterize inhibitory and excitatory neuronal cell types in the optic tectum, we use live imaging of electroporated single cells in Stage 47 Xenopus laevis and reconstructed the neurons' dendritic and axonal morphology. GABA immunolabeling of cross sections was used to determine whether the cell is inhibitory or excitatory. Here we present the comparison of excitatory and inhibitory neurons in the optic tectum based on their dendritic morphology and axon projections . Specifically, we compare the neurons' total dendrite length, number of branch tips, deepest dendrite depth, and axon projections, to gather whether any of these parameters may differentiate between excitatory and inhibitory neuronal cell types.
Abstract Title : Effects of invasive Foeniculum vulgare on pollination of neighboring Eriogonum arborescens
Abstract : Non-native plant species can disrupt plant–pollinator interactions by altering the foraging behavior pollinators and by contributing to the transfer of pollen to native plant species. In this study I investigate these potential effects in fennel (Foeniculum vulgare), which has become common throughout coastal California since its introduction to California from Europe in the 1800s. Fieldwork for this study took place on Santa Cruz Island, Santa Barbara County. I conducted a controlled and replicated fennel-flower removal experiment to test two hypotheses: (i) fennel flower removal alters visitation by insect pollinators on flowers of native plants, and (ii) fennel flower removal decreases the transfer of fennel pollen to flowers of native plants. The native plant considered in this study was the Santa Cruz Island buckwheat (Eriogunum arborescens). After the experimental removal of fennel flowers, insect visitation to buckwheat flowers was significantly greater in the control group than in the fennel flower removal group. Overlap in pollinator sharing between the two plant species was relatively low, but the buckwheat and fennel did share at least some pollinators. Native sweat bees (Agapostemon texanus) preferentially visited buckwheat, whereas yellowjacket wasps (Vespula pensylvanica) preferentially visited fennel. Fennel pollen transfer was unaffected by the removal of fennel flowers. These results indicate that fennel acts as a magnet species for pollinating insects and that this attraction spills over to native plant species. The results also indicate that insects readily transfer fennel pollen beyond the 13m radius of the experimental removal plots. Additional research could focus on the extent to which fennel pollen transfer hinders native plant reproduction.
Abstract Title : Preliminary study on the mechanism underlying α-syn impact on APP trafficking
Abstract : Parkinson’s Disease (PD) and Alzheimer’s Disease (AD) are two of the most common neurodegenerative diseases in the world. PD is typically characterized by pathogenic aggregates of α-synuclein (α-syn) that contribute to the loss of dopaminergic neurons in the substantia nigra par of the midbrain. AD, on the other hand, is manifested by the presence of β-amyloid plaques and tau containing neurofibrillary lesions in the brain. Intriguingly, many patients with PD also develop β-amyloid plaques, which indicates an overlap of the two diseases. To investigate the mechanism(s) of how pathogenic α-syn aggregates cross-talk with AD pathogenic pathways, we examined if and how α-syn in a mouse model of PD impacted trafficking and processing of amyloid precursor protein (APP) to promote production of Aβpeptide and pathogenic tau species. Consistent with previous studies showing that retrograde axonal transport is compromised significantly in PD neurons, we demonstrated that axonal trafficking of APP was selectively compromised in neurons from a transgenic PD mouse model that expresses GFP-α-syn. Because endosome-lysosome pathway is the main pathway that accounts for protein degradation, we looked at the endosome (rab5/rab7) and lysosome pathways that are the crucial loci where APP is cleaved by secretase. We found this pathway was widely impaired, with rab5-early endosome/ rab7-late endosome and lysosome trafficking damaged, which may account for accumulation of α-syn and APP byproducts in the pathogenesis of PD. Our observations thus far suggest that α-syn overexpression alters axonal trafficking of APP that likely results in enhanced intraneuronal production of Aβpeptides in PD neurons. Therefore, our study provides a plausible mechanistic insight into the molecular mechanisms that underlies AD-like dementia in PD patients.
Advisor : RU BRYAN
Abstract Title : Investigating the role of Adenosine Monophosphate-Activated Protein Kinase (AMPK) in a Murine Model of Obesity-Induced Osteoarthritis
Abstract : Osteoarthritis (OA) is a progressive and degenerative joint disease in which aging, obesity and joint injury are the major risk factors. OA is currently the most common joint disorder in the United States resulting in a breakdown of joint cartilage and bone causing stiffness and pain which affects approximately 10% of males and 13% of women over the age of 60. Risk of development is higher in individuals who are overweight or have high joint stress due to mechanical stress on joints which results in increased inflammation, however the underlying mechanism is unclear. Adenosine monophosphate-activated protein kinase (AMPK) is an essential regulator of energy metabolic homeostasis and is a factor in a variety of metabolic stresses as well as in the aging process. Studies have shown that AMPK signaling enhances energy metabolism and at the same time, can repress inflammatory responses linked to chronic stress. As AMPK declines in metabolic stress and aging it may cause metabolic diseases and accelerate the aging process. AMPK activity is observed in healthy human knee cartilage. It has been shown in previous studies that AMPK deficiency in chondrocytes accelerate the progression of OA in adult mice. We will investigate the role in obesity-induced onset OA via the role of AMPK.
Abstract Title : Analysis of hippocampal place cell dysfunction in mouse models of Alzheimer’s disease through single unit electrophysiology and calcium imaging techniques.
Abstract : Alzheimer’s disease (AD) is a neurodegenerative disease characterized by progressive mnemonic deficits. Previous studies have established that AD causes neural dysfunction in hippocampal place cells in the form of increased hippocampal place field size, decreased information score and decreased place cell stability in mouse models. Most such experiments, however, have been unable to record from specific layers of the hippocampus while mouse models complete goal-oriented spatial memory tasks. Here, we introduced a spatial memory task using a figure 8 scheme to allow electrophysiology recordings from the CA1, CA3, and DG regions of the hippocampus while young and aged transgenic AD mouse models (CA3-3xTg animals) completed the task. We also aimed to develop calcium imaging as a viable technique to analyze populations of place cells from the CA1 region at a larger scale relative to tetrode wire electrophysiological recordings, and we hoped to compare data retrieved using calcium imaging to data analyzed using tetrode wire electrophysiology. We found that CA1 and CA3 place cells indeed showed decreased information score, decreased trial by trial stability, increased spatial correlation, decreased place field size, and impairments in firing rates while animals completed memory tasks, particularly as the task became more hippocampal dependent. Animals tended to show more significant deficits in CA3 place cells relative to CA1. While our calcium imaging experiments were unsuccessful, we were able to set the foundation for future calcium imaging experiments in this work. Our results as a whole point towards substantial CA1 and CA3 place cell dysfunction while mouse models for AD complete spatial memory tasks as the task becomes more hippocampal dependent. Furthermore, our experiments set the stage for using calcium imaging as method that can add to previously existing information about place cell dysfunction in AD and therefore paint a more global image of hippocampal dysfunction in AD.
Abstract Title : Combining Stiffness and Stretch to Study Cardiac Fibroblast Pro-fibrotic Activity
Abstract : Cardiac fibrosis, defined by the excessive accumulation of the extra-cellular matrix, can cause severe cardiac problems and heart failure through stiffening the myocardial (heart) walls. Cardiac fibroblasts are the primary regulators of extracellular matrix production by increasing pro-fibrotic gene expression (col1a1, col1a2, & acta2) in response to mechanical stimuli. The mechanical stimuli, such as incessant stretching on the myocardium from chronic hypertension, initiates pro-fibrotic gene expression through mechanotransduction pathways. However, the details of how mechanotransduction pathways such as transforming growth factor β (TGFβ) and Rho kinase (ROCK) contribute to fibrosis and whether their functionalities differ in healthy versus fibrotic hearts remains to be clarified. We attempt to determine which mechanotranduction pathways are activated by stretching mouse cardiac fibroblasts plated at different stiffnesses representative of healthy and fibrotic hearts using in vitro stretching models. We observed elevated levels of col1a1, col1a2, and acta2 mRNA expression in cardiac fibroblasts representative of both a healthy and fibrotic hearts after being stretched. Our results further revealed that treatment with a TGFβ inhibitor before stretching the cardiac fibroblasts abolished any stretch-induced increase in mRNA expression. Treatment with ROCK inhibitor did not impact pro-fibrotic gene expression after stretching the cardiac fibroblasts. In conclusion, these findings indicate that the TGFβ pathway significantly contributes to fibrosis-related gene expression and that the TGFβ pathway functions are the same in cardiac fibroblasts from healthy and fibrotic hearts.
Abstract Title : The Importance of B-Cell Lymphoma 6 (BCL6) in Muscle Regeneration and Fat Metabolism
Abstract : B-Cell Lymphoma-6 (BCL-6) is a gene that encodes a transcription factor that represses genes necessary for the terminal differentiation of lymphocytes within germinal centers of B-cells. A novel drug, FX1, which binds to the BTB domain on BCL-6 was used in analyzing the important effects that this gene has on the proliferation and differentiation of myocytes. It was shown that FX1 inhibited the proliferation rate of myocytes in vitro and this drug was implemented to in vivo studies. These findings also indicated that there was a decrease in fat mass while maintaining lean muscle mass.
Abstract Title : Evolutionary Consequences of CRISPR/Cas9-based Gene Drives in S. cerevisiae
Abstract : CRISPR/Cas9-based gene drive technologies are poised to solve current struggles against tropical, vector-borne diseases such as Zika, dengue, and malaria. Once introduced into a population, many gene-drive constructs will remain fixed in the population. We hypothesized that the long-term presence of an active gene-drive element may affect the evolution of that population. To test this hypothesis, we took a two-pronged approach. In the first experiment, we evolved populations of diploid baker’s yeast, Saccharomyces cerevisiae, expressing a full Cas9-based gene drive system (i.e., Cas9 + gRNA), only Cas9 without a gRNA, or a control expressing neither Cas9 nor gRNA. By tracking the frequencies of new and pre-existing mutations across 8 replicate populations per strain over 1000 generations, we observed signatures of adaptation and estimated the rate at which new mutations arise and fix in populations from each treatment. In the second experiment, we measured the relative mutation rates and rates of gene conversion events in the same three treatments. To this end, we carried out a mutation accumulation assay for about 600 to 700 generations per replicate line. In our initial analyses of the asexual evolution experiment, we observe that more de novo mutations reach high frequencies in populations with Cas9 with an especially pronounced effect in the strain lacking a gRNA. We are currently processing samples from the mutation accumulation experiment for DNA sequencing. The resulting mutation rate estimates from this assay will confirm whether Cas9 nuclease is mutagenic and/or elevates gene-conversion rates in our strains. Our results will inform us on the long-term safety of CRISPR/Cas9-based gene drives.
Advisor : MILTON H. SAIER, JR.
Abstract Title : Structural bioinformatic characterization of the tetraspanning junctional complex (4JC) superfamily
Abstract : Intercellular connections are critical to the development and health of multicellular eukaryotes. These connections link neighboring cells with each other, adjacent extracellular spaces, and organelles and the cytoplasm and are typically mediated by membrane proteins, many of which are capable of sensing changes in membrane shape contributing to hearing and mechanotransduction, transporting ions or other substrates, or relaying a variety of other signals. The tetraspanning junctional complex (4JC) superfamily comprises proteins found in tight junctions, gap junctions, mechanosensitive cells, and various ion channels. Many of the families proposed to be members of the 4JC superfamily exhibit low sequence similarity with each other, confounding past attempts at building a case for homology. Nowhere is this more evident than in the innexins, gap junction proteins for invertebrates once posited to be entirely unrelated to the connexins, gap junction proteins for vertebrates. Thanks to the resolution revolution in electron cryomicroscopy and recent boom in membrane protein structures, however, we are now able to reproduce various family-family comparisons in structure as well as in sequence. We anticipate that these findings will facilitate attempts at predicting interaction partners for 4JC proteins and elucidating the development of multicellularity.
Abstract Title : MicroRNAs Associate with the Caenorhabditis elegans Argonautes ALG-1 and ALG-2 to Differentially Regulate Longevity
Abstract : MicroRNAs (miRNAs) are small non-coding RNAs which negatively regulate gene expression through the binding of an Argonaute (AGO) protein which facilitates the translational repression or complete degradation of a target messenger RNA (mRNA). MiRNAs were first discovered in the nematode Caenorhabditis elegans, with Argonaute-like Gene 1 (ALG-1) and ALG-2 acting as the main AGOs in the miRNA-silencing pathway. Previous work has shown ALG-1 and ALG-2 function similarly during development but alg-1 and alg-2 loss of function (LOF) during adulthood has shown to produce opposing lifespan phenotypes; with alg-1 and alg-2 mutants during adulthood reducing or increasing longevity respectively. The divergent lifespans observed in alg-1 and alg-2 mutants were shown to cause the misregulation of several aging-associated miRNAs, such as lin-4 and miR-239a/b, which differentially regulate longevity through the IGF/Insulin signaling (IIS) pathway. Although several miRNAs have already been implicated in causing these divergent lifespans, the full extent of miRNA involvement has yet to be illuminated. SmallRNA profiling performed on these mutants during adulthood identified 51 and 19 significantly misregulated miRNAs in alg-1 and alg-2 backgrounds respectively. MiRNA-mediated regulation of aging-associated pathways is a highly conserved phenomenon in most eukaryotes, including humans. By understanding how perturbation of ALG-1 and ALG-2 differentially regulates aging in C. elegans, we can increase our understanding of the molecular basis of aging as a whole.
Abstract Title : Generation and characterization of a Shpk knockout mouse model and impact on HSC transplantation therapy for cystinosis
Abstract : Cystinosis is a lysosomal storage disease caused by the deletion of CTNS gene. The most common mutation in patients also partially eliminates sedoheptulose kinase (SHPK), a metabolic enzyme involved in modulation of macrophage polarization. Previous studies revealed that hematopoietic stem cell (HSC) transplantation into Ctns-/- mice prevents disease progression via HSC-derived macrophage-mediated delivery of functional lysosomes to diseased cells. We, therefore, analyzed the impact of Shpk elimination on this HSC therapy. We CRISPR-generated and characterized the first Shpk-/- mouse model and analyzed HSC-transplantation efficacy in Ctns-/- mice. This research is critical for elucidating which patients can benefit from upcoming HSC-gene therapy clinical trials.
Abstract Title : The effect of PHLPP2 removal on cardiac ischemia/reperfusion (I/R) injury and signaling in vivo
Abstract : The serine/threonine phosphatases, PHLPP1 and PHLPP2 (PH domain Leucine-rich repeat protein phosphatase) attenuate the activity of Akt via dephosphorylation in several cell types. Little is known regarding the biological function of PHLPP isoforms in the heart or its role in regulating pathophysiological stress. We previously found that phlpp1 null mice demonstrated increased Akt activity and phosphorylation at Ser473 in the heart at baseline, which conferred protection following ischemic damage (Moc C et al. 2015). We have recently generated phlpp2 null mice; however, preliminary studies demonstrate no significant changes in Akt phosphorylation in the hearts at baseline. It is important to understand whether the isoforms have specific or redundant functions in the heart. In this study we wanted to determine the effect of PHLPP2 removal on cardiac ischemia/reperfusion (I/R) injury and signaling in vivo. Wild-type (WT) and PHLPP2 knockout (KO) mice were subjected to 1hr ischemia and 24hrs reperfusion. We determined that PHLPP2 removal accentuated the percent infarct relative to area at risk (40.4 ± 2.2%, n=6) compared to WT (33.3 ± 1.5%, n=6) using triphenyltetrazolium chloride (TTC) and Evan’s blue dye. Ischemic, border and remote zone of hearts following I/R will be examined by mRNA analysis to elucidate the pathways altered by PHLPP2 removal. Our findings demonstrate for the first time that PHLPP1 and PHLPP2 do not have redundant functions in the heart. Further studies will determine the signaling pathways altered by PHLPP removal in the heart to identify new therapeutic targets for heart disease.
Abstract Title : Pedagogical Miscommunication in Biological Symbolic Representation: Arrows
Abstract : Biology frequently describes many abstract phenomena such as, but not limited to, DNA replication in the cell growth cycle, proton transportation in the energy production process, and ligand activation in the immune response system. Instructors often communicate such complex phenomena through symbolic representation. Student performance on exams and research papers can gauge limited understanding of the concepts, but may not identify the variations in the conceptions of the fundamental elements onto which we want students to build a solid understanding of abstract ideas. For example, it is uncommon for instructors to have time or opportunity to examine how well students understand some of the essential aspects of biology representations. Since the student population in the university system is becoming more diverse, we wanted to examine the relationship between English familiarity and interpreting one of the most common symbols used in biology textbooks, arrows. Through surveying 1969 students in multiple introductory biology courses, we evaluate the preciseness and consistency of students' understanding of various types of arrows in the biological context. In our findings, English proficiency is correlated with students' decision when decoding the meaning of an arrow. In our exploratory research on the possible relationship between the visual representation and verbal description of arrows in biology, there exists enormous variation in understanding the meaning of arrows among all students, regardless of language status. We suggest the instructors in the biological field restate and enforce the specific and consistent usage of symbols.
Abstract Title : Elevated Erucic Acid Concentration in Women with Anorexia Nervosa
Abstract : Background: Anorexia nervosa is the deadliest eating disorder and psychiatric illness whose etiology remains unclear. Anorexia nervosa patients have shown an aversion toward foods high in fats and calories, which may have hindered treatment success as suggested by lower fat intake in anorexia nervosa patients with poor treatment outcomes when compared to patients with successful treatment outcomes. The high-fat foods that anorexia patients avoid are one of the major sources of omega fatty acids, which play important roles in healthy brain development and are implicated in cardiovascular, inflammatory, and psychiatric disorders. Additionally, altered levels of omega fatty acids have been found in patients with eating disorders, suggesting possible involvement in the pathophysiology of eating disorders. Thus, this study aims to explore the role of common omega fatty acids in the risk and clinical phenotypes of anorexia nervosa. Methods: Blood samples were collected from women with anorexia nervosa and age- and sex-matched healthy controls before and two hours after the subjects ate a breakfast sandwich (fasting N= 6 anorexia nervosa patients and 16 controls; postprandial N=6 anorexia nervosa patients and 12 controls). Questionnaires were administered to all study subjects to assess anxiety (BAI), depression (BDI), and food aversion (FAQ). Additionally, anorexia nervosa patients completed two eating disorder questionnaires (EDE-Q and EDI). Mass-spectrometry based lipid analysis was used to quantify both fasting and postprandial plasma levels of seven common fatty acids, which included two omega-6, four omega-3, and one omega-9 fatty acid. Results: Of the 7 fatty acids, most of them showed no statistically significant difference between women with anorexia nervosa and healthy controls, both at fasting and postprandial timepoints. Fasting levels of the monounsatured omega-9 erucic acid were significantly higher in anorexia nervosa patients than in healthy controls (136.95 ±75.29 pmol/ml and 77.16 ±67.68 pmol/ml, respectively; p=0.01). Similarly, in the fed state, women with anorexia nervosa had higher plasma levels of erucic acid than controls (124.18± 49.61 pmol/ml and 67.05±33.51pmol/ml, respectively; p=0.02). At the fasting timepoint, women with anorexia nervosa showed a positive correlation between erucic acid and change in BAI (rho=0.90, p=0.04), whereas controls exhibited a negative correlation between erucic acid and change in BAI (rho= -0.83, p=0.04). Erucic acid was not significantly correlated with BMI, BDI, or specific food aversions in both patients and controls. Conclusion: Compared to healthy controls, women with anorexia nervosa showed significantly higher levels of erucic acid both before and after eating a sandwich enriched with omega-6 arachidonic acid, suggesting that erucic acid may be involved in the pathophysiology of anorexia nervosa. However, further studies with larger sample sizes are needed to confirm the association between erucic acid and anorexia nervosa status and the (lack of) correlations between erucic acid levels and comorbid phenotypes in anorexia nervosa.
Abstract Title : Synapse formation, processing, and retention is altered in Huntington’s mice cortical neurons.
Abstract : Huntington’s (HD) is a genetic disease caused by an expansion in the poly-Q region of the Huntington gene resulting in the preferential death of dopaminergic neurons in the striatum. Death of the dopaminergic neurons results in the motor symptoms seen in the early stages of the disease and it’s currently unclear why these neurons are the most vulnerable. Using mouse models, we can try to understand the cellular and molecular mechanisms that underlie the death of these neurons. Previous research has suggested brain derived neurotrophic factor (BDNF) release by cortical neurons as well as BDNF signaling through p75 and TrkB in striatal neurons might be disrupted. Work has also shown that BDNF transport is impacted in cortical neurons which can be rescued using apical chaperonin containing TCP-1 (ApiCCT1). Additionally, it is believed that excitotoxicity resulting from excess glutamate release from cortical neurons may play a role in striatal neuron death. Here, we aim to further explore how HD cortical neurons might be disrupted in mass culture by quantifying synapse formation, processing and retention. HD cortical neurons showed dramatically different synapse processing with a verity of phenotypes potentially related to synaptic protein trafficking. Additionally, we try to rescue deficits seen in HD cortical neurons using ApiCCT1 which has previously been shown to rescue transport in HD neurons.
Abstract Title : The Expression of Downregulated in Adenoma (DRA) After Salmonella Infection and the Inhibition of NF-kB
Abstract : Salmonella has been identified as a foodborne illness that is responsible for over one million hospitalizations yearly in the United States alone. Specifically, Salmonella disturbs the overall function of the intestinal epithelium and its ability to regulate the absorption process of electrolytes. Downregulated in Adenoma (DRA) is a transporter protein in the intestinal epithelium that regulates the sodium chloride absorption balance and has been found to be downregulated upon Salmonella infection. Other pathways like the NF-kB pathway have also been found to also downregulate DRA expression as well. However, the mechanism to which Salmonella downregulates DRA expression is still unknown and is thought to possibly act on this particular pathway. Therefore, by inhibiting the NF-kB pathway, this can serve as a potential therapeutic to upregulating DRA expression and to ultimately combat Salmonella infection.
Advisor : ANJANA RAO
Abstract Title : Investigating the role of NFAT2 Transcription Factor in T Cell Exhaustion
Abstract : T cell exhaustion is a state of dysfunction caused due to persistent antigen exposure that occurs in cancer and chronic infections. It results in loss of ability to perform effector functions and is characterized by sustained expression of inhibitory receptors, diminished cytokine production and decreased survival. Our lab has previously demonstrated that NFAT1, a member of the NFAT transcription family, plays a pivotal role in controlling the program of T cell exhaustion. However, the role of other NFAT transcription factors is not known. In this study, we seek to define how NFAT2 contributes to exhaustion, and to determine whether NFAT2 and its individual protein isoforms have distinct roles in overall transcriptional program of T cell exhaustion by using Flowcytometry, RNA-Seq and ATAC-Seq.
Abstract Title : Characterizing the Function of VICTR-like Genes and Their Role in the Regulation of ABA in Response to DFPM in Arabidopsis Plants
Abstract : The hormone abscisic acid (ABA) triggers signal transduction that mediates drought resistance. A synthetic small molecule “DFPM” ([5-(3,4-dichlorophenyl) furan-2-yl]-piperidine-1- ylmethanethione) has been shown to negatively regulate ABA signaling and stimulate plant defense related genes. The Variation In Compound Triggered Root growth response (VICTR) locus is required for DFPM-mediated root growth arrest in Arabidopsis thaliana. VICTR encodes a Toll-Interleukin1 Receptor– nucleotide binding–Leucine-rich repeat (TIR-NB-NLR) protein. To determine whether VICTR and its homologous tandem genes play a role in DFPM inhibition of ABA signal transduction we utilized a Near Isogenic Line (NIL) NIL-Bu-5 of Arabidopsis with accessions of the Bu-5 ecotype crossed into the background of Columbia-0 (Col-0). NIL-Col-0 which contains functional VICTR and its homologous genes and NIL-Bu-5 which lacks all four tandem VICTR homologs were used in this study. We investigated whether VICTR and the VICTR-like genes contribute to the regulation of ABA signal transduction. We first grew seedlings of Col-0, NIL-Col-0, and NIL-Bu-5 for two weeks, exposed plants to various chemical conditions including ABA, DFPM, and solvent control for 5 hours followed by RNA extraction, complementary DNA synthesis, and reverse transcriptase-quantitative PCR to analyze expression of ABA reporter genes RAB18, RD29A, and ERD10. We analyzed fluorescence intensity of the pRAB18::GFP ABA signaling reporter after chemical treatments and compared Mitogen-Activated Protein Kinase (MAP Kinase) activity between genotypes after DFPM treatment under various exposure times. DFPM-mediated MAP Kinase activation was not disrupted when VICTR and its homologs are lacking. The roles of VICTR and its homologs are being studied with respect to ABA signal transduction under ABA and DFPM chemical treatments, further analysis will allow us to understand the function of VICTR and its homologous genes in DFPM signal transduction.
Abstract Title : Exploring the Origin of Eukaryotes: Comparative Genomics of Transport Systems between the Asgard Archaeal Superphylum and the Tree of Life
Abstract : Upon discovery of the first archaeal cell in the 1970s, life has been differentiated into three domains: Eukarya, Archaea, and Bacteria. However, the organization of the three-domain tree of life has been challenged following the discovery of the archaeal lineages such as the superphyla TACK and, most recently, the Asgard. The Asgard super phylum has emerged as the closest predecessor to the eukaryotes and may improve our understanding of the evolution of life from relatively simple prokaryotes to complex eukaryotes. Here we characterize the transportomes of four metagenomes of Asgard archaea (i.e. Loki-, Odin-, Thor-, and Heimdall-archaeota). Candidate transporters were identified based on their similarity to transporters within the Transporter Classification Database (TCDB), and shared hydropathy profiles, TMS topologies and Pfam Domain contents. Identified transport systems will be compared within the Asgard superphylum, and to other eukaryotic, archaeal, and bacterial transport systems. This comparison should provide insight into the physiological similarities and differences between Asgard genomes and other characterized organisms (cultured and uncultured). Furthermore, this analysis may provide insight into the hypothesis suggesting an archaeal origin of eukaryotes. The identification of transporters unique to eukaryotes in the Asgard superphylum would suggest that components of eukaryotic complexity arose before the emergence of the first eukaryotic cell.
Abstract Title : Exploring the Temporal and Spatial Development of Neuronal Cell Types in the Tadpole Brain
Abstract : The neurons of the central nervous system are grouped into distinct classes; historically, this classification has largely been based upon neuron structure and function. However, genetic differences between neuronal subtypes are being increasingly utilized as a measure of distinction between prospective neuron classifications. In particular, differential expression analysis has been successfully used to establish neuronal cell types based on their distinct profiles of RNA expression. Such studies help illuminate the wide diversity of neurons present in the vertebrate nervous system and provide researchers with a foundation on which to base further investigations in the field. As a ubiquitous model for such neurobiological research, gene expression in the developing tadpole Xenopus laevis (the African clawed toad) has been well-documented in literature. We sought to quantify gene expression in the X. laevis brain in two ways: first, over consecutive stages of development, and second, across distinct regions of the brain (the fore-, mid-, and hindbrain regions). We then compared lists of genes we found to be differentially expressed between developmental stages, as well as between brain regions, against lists of genes curated from the literature that were found to be specific to certain neuron subtypes. We hypothesize that significant enrichment of such neuron class-specific genes may suggest a time interval or brain region in which these classes develop. Utilizing both temporal comparisons across development as well as spatial comparisons across brain regions, then, we will illuminate the chronology and locality in which neuron classes arise in the developing Xenopus brain.
Abstract Title : Adaptive laboratory evolution of Carnobacterium sp. AT7 under increased hydrostatic pressure
Abstract : The ocean makes up over 97% of the Earth’s biosphere and, with an average depth of 3,800 meters, much of it is located within deep-sea habitats. Life at these depths must possess adaptations for growth at high hydrostatic pressures. Microorganism that grow preferentially at elevated pressures are termed “piezophiles.” Although prior studies have examined the adaptations of piezophilic and piezotolerant Gram-negative microbes, little attention has been given to Gram-positive bacteria. In this project, we examined the only-known Gram-positive piezophile, Carnobacterium sp. AT7. Two separate lineages of AT7 were subjected to progressively increasing hydrostatic pressures over the course of 100 generations. From these lineages, we obtained mutant strains which possessed improved rates of growth at high pressure and increased upper pressure limits. During the course of these experiments, it was discovered that incubation of AT7 under stressful, high hydrostatic pressure for a period of approximately 200 hours resulted in the acquisition of greatly improved high pressure growth in a single incubation cycle. Interestingly, cells derived from these incubations did not display stable high pressure growth and are presumed to have gained this ability through a physiological mechanism, rather than a genetic one. Because it is well known that high pressure growth requires the production of sufficient unsaturated fatty acids, we are investigating whether the mutant strains, or long-term incubation cultures, display increased proportions of unsaturated fatty acids. Possible morphological alterations are also being assessed. The genetic basis of improved piezophily in the mutant strains will be determined by genomic re-sequencing. The results obtained will shed light on the processes used by Gram-positive bacteria to adapt to elevated high pressure conditions.
Abstract Title : An integrated RNA-Seq and proteomics profile of stem tissue following Fusarium venenatum elicitation reveals major patterns of gene expression and provides examples of translational programming in maize
Abstract : Maize is a major staple food crop with important agricultural and agronomic impact worldwide. However, fungi, including those of the genus Fusarium, can cause serious diseases in maize, resulting in significant economic losses. To investigate how fungal challenge affects maize, we slit the stems of maize plants and elicited a defense response with heat-killed Fusarium venenatum hyphae. We then collected stem tissue over a ten-point time course and obtained RNA-seq and proteomics fold change data compared to the 0 hour timepoint. Our findings indicate that the maize transcriptional and translational responses to fungal elicitation do not necessarily correspond to each other. Weighted gene co-expression network analysis (WGCNA) resulted in a module in which RNA does not change over the time course but protein abundance increases, providing examples of translational programming in maize. GO analysis on the genes in this module reveals enrichment for protein modifications and vesicle-mediated transport. Performing WGCNA on the fold change rank orders yields four main patterns of RNA and protein expression. In the largest rank-order module, RNA and protein abundance both decrease over time; this module is most enriched for genes involved in plant development, providing evidence for the shift between growth and defense. The next three largest modules exhibit increasing protein abundances, and are enriched for genes involved in ubiquitination and responses to stimuli, which are all involved in defense. We hope that our transcriptional and proteomic analyses will lead to a better understanding of plant immunity and ultimately reduce economic losses caused by fungal infection.
Abstract Title : A Heterologous Phosphite Oxidation Gene ptxD Enables Cyanobacteria to Utilize Phosphite as Sole Phosphorus Source
Abstract : Phosphorus is an essential nutrient for all organisms. Many obtain phosphorus in the form of phosphate, which is readily available in the environment. A few microbes are capable of utilizing other reduced forms, including phosphite and hypophosphite, using a phosphite-oxidative enzyme that converts them to the usable phosphate. Due to the limited number of organisms that can utilize phosphite, restricting the sole source of phosphorus to phosphite has applications in culturing cyanobacteria. For example, growth of contaminating competitor strains can be reduced in industrial large-scale algae ponds and isolation of genetically tractable wild strains from environmental samples could be facilitated by transformation with genes for phosphite utilization. In order to confer cyanobacterial strains with the ability to oxidize phosphite, the ptxD gene encoding phosphite oxidoreductase was extracted from Pseudomonas stutzeri and assembled into an RSF1010-based broad-host range plasmid. Three cyanobacterial strains were transformed with a plasmid containing the ptxD gene: Anabaena PCC7120, Synechocystis PCC6803, and Leptolyngbya BL0902. Our results show that the strains are able to grow in cyanobacterial BG-11 media with phosphite substituted for phosphate, and growth was affected by the concentration of phosphite.
Abstract Title : Identifying Genetic Players in Stomatal Drought Signal Repression and Pathogen Infection in Arabidopsis Plants
Abstract : Being sessile organisms, plants have evolutionarily developed a variety of mechanisms to protect themselves from biotic and abiotic stressors such as pathogen exposure, drought conditions, and heavy metal contamination. The phytohormone, abscisic acid (ABA), is a major stress hormone in Arabidopsis thaliana essential to resistance of abiotic stressors, such as drought conditions. In addition to mediating drought tolerance, however, ABA has been found to interfere with pathogen resistance signaling as well by increasing susceptibility to pathogens. Recent discoveries demonstrated a novel interference in plant stress signaling-- ABA signal repression by pathogen defense activation. In a chemical genetics approach it was found that exposure to a newly identified small molecule, [5-(3,4-dichlorophenyl)furan-2-yl]-piperidine-1-ylmethanethione (DFPM), down-regulates ABA-signaling via pathogen resistance signaling pathways in plants. While this exposure-induced phenotype is consistently evident in A. thaliana accession Col-0, the molecular and cellular mechanisms of this pathogen signaling repression of drought signaling are poorly understood. Through a forward chemical genetics screen, rda mutants (resistant to DFPM inhibition of ABA signaling) were identified. Our study eliminates gene loci predicted to be the causative mutation of this rda mutant and identifies transcription factors required for the DFPM mediated immune signaling.
Abstract Title : The Effect of Project Ownership on Student Approaches to Writing in Laboratory Courses
Abstract : Course-based undergraduate research experiences offer not only the opportunity to engage students in aspects of research, but also other important scientific thinking and process skills such as written communication. Engaging in authentic scientific writing would involve writing from a place of understanding (a “top-down” approach) rather than relying heavily on given ideas and information to dictate how to write and what is written (a “bottom-up” approach). Indeed, students’ perceptions of authorial identity have previously been shown to vary and be affected by course structure (such as rubrics) and the approaches they take to writing. Our previous work suggests that approaches to writing may be influenced by the sense of ownership students have over a project. Given the connectedness of research ownership and authorial identity, we wanted to explore the relationship between ownership and approaches to writing. Although previous research has explored how student descriptions of their research experiences can reveal concepts of ownership, we do not have a clear idea of how students perceive ownership in a lab class setting. In addition, we do not have a clear idea of how approaches to writing are related to perceptions of ownership, particularly in a course-based research setting. This led to our current questions: 1) How do students perceive ownership in a lab course? 2) How does an increase in project ownership affect students’ approaches to scientific writing? To address these questions, we used qualitative analysis of student responses to questions administered as a survey, and during interviews. Students were surveyed from a specific upper-division laboratory course where they engage in multiple lab modules, one of which is considered to have a low level of inquiry (little choice over design, predictable outcomes, and they are not addressing a sophisticated research question), and another module which has greater inquiry (students have choice over the experimental design, the outcomes of their experiment are unknown to the students and the instructor, and the research question is more sophisticated). Students write short journal article-style reports about each project. Students were provided with an opportunity to take a survey or participate in an interview, both of which asked various questions about how they defined ownership, their sense of ownership over the lab projects, and how they approached their writing for the two projects. Using a grounded theory approach for the analysis of 140 survey responses and 13 interviews, we found that students have varying perceptions of ownership, despite working on the same projects. Common themes that emerged including ownership meant contributing ideas, doing the work, being responsible for outcomes, as well as relationships between ownership and understanding of the project. Student approaches to writing were largely considered “bottom up” as opposed to the “top down” approaches associated with a greater sense of authorial identity. These findings provide insights about how course design can impact not only a student sense of ownership, but how this sense of ownership influences the type of scientific writing they do. In particular, despite efforts to increase ownership, other course structures may reduce the sense of authorship students have and demonstrate. If our goal is to provide research, including writing experiences through a course-based laboratory setting, we need to carefully consider how scientific writing is incorporated, as well as how course structure supports the development of a sense of ownership and authorship.
Advisor : DR. YISHI JIN
Abstract Title : Regulatory role of translation initiation factor EIF-3.G in Caenorhabditis elegans neural circuit.
Abstract : Regulated neural circuit activity is essential for normal brain function whereas aberrant circuit function is a hallmark of neurological disorders, including Autism Spectrum Disorders and epilepsy. The nematode C. elegans has been a prime animal model for investigating genetic mechanisms that regulate neural circuits. Its simple neural anatomy and powerful genetic analysis enable studies to be carried out with precise resolution. Moreover, many genes that are first discovered in C. elegans have similar roles in mammals. Here, we examine neuronal circuit regulation using a gain-of-function mutation in a nicotinic acetylcholine receptor subunit, acr-2(gf), which hyperactivates the cholinergic motor neurons, causing impaired locomotion and stochastic muscle convulsions. Through genetic screening, we have identified a functional pathway linking neuronal activity with the conserved translation initiation factor eif-3.g. EIF3 is a multimeric eukaryotic initiation factor with an essential role in initiation of protein translation. EIF-3.G is one of four subunits that contain a known RNA-binding domain. We employed a genetic suppressor screen to reveal the genetic pathway affecting motor neuron function through eif-3.g. Here, we describe approaches used to map causative mutations identified from this screen. Based on strain-specific SNPs revealed by whole genome sequencing analysis, we traced SNPs among many recombinant strains. We have so far identified two eif-3.g suppressors, the protein translation repressor lin-66 and a predicted protein phosphatase eat-9. We speculate these genes are targets of or participate in eif-3.g function in motor neurons. Our studies provide fundamental mechanism for future implication on neurological comprehension and therapeutics of neural circuit related disease.
Advisor : STANLEY LO
Abstract Title : Examining the variations in undergraduates’ conceptions of researchers: A phenomenographic study
Abstract : Undergraduate research experiences play a critical role in training the next generation of scientists; these experiences also increase students’ persistence in science and enhance their cognitive and personal development. However, participation in research requires awareness of existing opportunities and an understanding of the cultural norms within the research community. While some studies have identified the various ways in which researchers understand their profession, few studies have directly examined student conceptions of researchers, and even fewer focused on undergraduates. Given that the norms and values of the research community may be unfamiliar to students, especially for first-generation (FG) college students who lack access to the cultural knowledge of research communities, our study examined how undergraduates experience or conceptualize researchers as a profession, using phenomenography as the theoretical framework. Phenomenography describes the qualitatively different ways that individuals experience, understand, or think about the same phenomenon. Within phenomenography, variation theory formalizes the ways of experiencing a phenomenon into an outcome space with specific features (called “aspects”) that describe the phenomenon, as well as the “variations” within these aspects that define the experiences. Data were collected from a summer-bridge program aimed at introducing incoming transfer students, who were disproportionately FG (75%), to biological research. To capture a wide range of research experiences, 29 participants were interviewed about two years after the program, near their target graduation date. Participants were asked to give their perception of what a successful researcher is and what a researcher specifically does to be successful. Participants were also prompted to indicate whether their conception of researchers had changed and what specific experiences prompted these changes. Data were analyzed through iterative close reading of the interview transcripts using grounded theory, which is divided into three major phases of qualitative coding. In open coding, memos and in vivo codes were generated using participants' own phrases that capture critical meanings of a researcher. In axial coding, specific aspects and variations were identified and organized into an outcome space that describes different conceptions of researchers. In selective coding, data for each aspect and variation were revisited to further refine the definitions in the outcome space. Three conceptions of researchers (Type I, Type II, and Type III) were identified based on variations within the following aspects: research process, interactions with other researchers and broader communities, and scope of contribution. Most participants retrospectively described their initial conceptions of a researcher as simplistic, with little appreciation for the complex methodological processes and extensive collaboration needed to meaningfully contribute to the research community (Type I). However, extensive research experiences had shifted their conceptions. Participants who changed to the Type II conception recognize research as a defined process that requires extensive collaboration and contribution within the research community. The Type III conception includes and expands upon Type II by viewing research as not only a defined but also iterative process that is used to interact with and contribute to both the research community and a broader society. The disconnection between the initially simplistic conception of researchers (Type I) and the more mature conceptions (Types II and III) indicates the conflicting expectations that students with limited exposure to research may have prior to their own research experiences. Overall, our findings support the need for broad-scale institutional interventions that engage students in research and provide guidance for transition into research.