Cell Biology & Molecular Genetics Theses and Dissertations
Permanent URI for this collectionhttp://hdl.handle.net/1903/2750
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Item Cell Population Shifts and Clinical Heterogeneity in Sjögren's Disease(2024) Pranzatelli, Thomas J; Johnson, Philip L.F.; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Sjögren's disease (SjD) is a systemic autoimmune disease that causes loss of function of the salivary and lacrimal glands. Those with the disease, overwhelmingly female with an onset of disease in the fourth or fifth decade of life, commonly suffer from dry mouth, cavities and damage to the eyes. Patients present with a wide variety of clinical phenotypes, with variation in degree of immune infiltration and glandular damage as well as positivity for autoantibodies. This thesis uncovers the changes in cell population and gene expression in the gland that underpin diversity in disease severity. SjD patients lose the majority of a specific epithelial population in their labial salivary glands and, as the number of immune infiltrates grows the surviving members of this population can be found colocalizing with invading GZMK+ T cells and expressing markers of increased proliferation. Standard differential gene expression analysis highlighted gene markers of cell types changing in proportion with disease; an unenlightening result when the cell population changes are well-characterized. To avoid this pitfall an ensemble of random forests was trained to find genes predictive of patient subtypes without being correlated with diagnosis. Genes with high importance for autoantibody positivity were enriched for GO terms related to antigen processing and presentation. A master regulator of salivary gland identity, ZBTB7B, was identified from chromatin accessibility data. Mice with this transcription factor knocked out lose salivary flow and develop pockets of tissue in their glands that resemble other glands, eg., labial gland epithelium inside of parotid glands. This work supports a clinical presentation-specific approach to therapy and paves the path for reengineering the glands to correct the effects of disease.Item DEVELOPMENT AND OPTIMIZATION OF TOOLS FOR CO-EXPRESSION NETWORK ANALYSES OF HOST-PATHOGEN SYSTEMS(2017) Hughitt, Vincent Keith; El-Sayed, Najib M; Cell Biology & Molecular Genetics; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)High-throughput transcriptomics has provided a powerful new approach for studying host-pathogen interactions. While popular techniques such as differential expression and gene set enrichment analysis can yield informative results, they do not always make full use of information available in multi-condition experiments. Co-expression networks provide a novel way of analyzing these datasets which can lead to new discoveries that are not readily detectable using the more popular approaches. While significant work has been done in recent years on the construction of coexpression networks, less is known about how to measure the quality of such networks. Here, I describe an approach for evaluating the quality of a co-expression network, based on enrichment of biological function across the network. The approach is used to measure the influence of various data transformations and algorithmic parameters on the resulting network quality, leading to several unexpected findings regarding commonly-used techniques, as well as to the development of a novel similarity metric used to assess the degree of co-expression between two genes. Next, I describe a simple approach for aggregating information across multiple network parameterizations, in order to arrive at a robust “consensus” co-expression network. This approach is used to generate independent host and parasite networks for two host-trypanosomatid transcriptomics datasets, resulting in the detection of both previously known disease pathways and novel gene networks potentially related to infection. Finally, a differential network analysis approach is developed and used to explore the impact of infection on the host co-expression network, and to elucidate shared transcriptional signatures of infection by different intracellular pathogens. The approaches developed in this work provide a powerful set of tools and techniques for the rigorous generation and evaluation of co-expression networks, and have significant implications for co-expression network-based research. The application of these approaches to several host-pathogen systems demonstrates their utility for host-pathogen transcriptomics research, and has resulted in the creation of a number of valuable resources for understanding systems-levels processes that occur during the process of infection.Item Evolution of sex-biased expression in Caenorhabditis(2011) Thomas, Cristel Gwenola; Haag, Eric S; Molecular and Cell Biology; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Mating systems have a profound impact on genome structure evolution, both indirectly through their effects on population genetics and directly due to the genetic control of reproductive traits. Most extant Caenorhabditis species are gonochoristic (males and females), while the most studied species, C. elegans and C. briggsae, are androdioecious (self-fertile hermaphrodites and males). The latter two species display an overall reduced ability to mate, suggesting that the selective pressure on maintaining efficient mating was weakened as selfing arose. The genes underlying these traits were likely to have been expressed in a sex-biased fashion in the gonochoristic ancestor, and we hypothesized that as selfing emerged their regulation was modified or they were lost altogether. This hypothesis is especially interesting given that selfing species have consistently smaller genome sizes than their gonochoristic relatives. I sought to address whether a disproportionate loss of genes with sex-biased expression accompanies the loss of mating-related traits in Caenorhabditis hermaphrodites. I first examine sex-biased expression in a gonochoristic species, C. remanei, and identify genes with highly sex-biased expression. I find that these genes are more likely to be missing in selfing species than expected by chance. I then select some of these genes based on their phylogenetic conservation patterns in the genus, and characterize them more thoroughly to shed some light on their functions. Through this study I identify a novel male-associated candidate cis-regulatory element. Lastly, I broaden the scope of the study by determining transcriptome wide sex-biased expression patterns in four Caenorhabditis species. I confirm that C.elegans displays a decrease in the proportion of strong female-biased expression, as well as a modification of the expression of genes with male-biased expression both in males and in hermaphrodites, when compared to gonochoristic Caenorhabditis. Taken together, this study illustrates the transcriptomic consequences of a modification of the mating system, and begins to address its effect on genome structure.Item Identification of a single nucleotide polymorphism associated with adiposity following transcriptional profiling of gene expression in the anterior pituitary gland(2006-07-23) muchow, michael; Porter, Tom E; Molecular and Cell Biology; Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)Although the anterior pituitary secretes three hormones that affect metabolism and body fat stores, a comprehensive analysis of pituitary gene expression associated with body fat has not been performed. This research used cDNA microarrays to investigate pituitary gene expression in two chicken lines that were selected for low and high body fat (Lean and Fat). RNA was extracted from pituitaries at 1, 3, 5, and 7 weeks of age. 386 genes that showed significant differences in expression levels by line or in the line-by-age interaction were analyzed further. Differentially expressed genes between lines are potential candidates as genetic markers for high and low potential for body fat accumulation. One such candidate, the lysophosphatidic acid (LPA) receptor-1 (LPAR1), was identified as a potential marker, being differentially expressed between the 2 lines at the early ages. Genomic DNA from the Fat and Lean F0 generation was sequenced upstream of the LPAR1 coding region. A SNP consisting of a T to C transversion that introduces a GATA-1 transcription factor binding site was identified in the Lean line (Fisher's Exact Test, p ≤ 0.001). The fattest and leanest animals of both sexes in the back-crossed F2 generation (n=48 each) were genotyped by allele-specific PCR, and an association was present between the genotype and phenotype (generalized linear model, p ≤ 0.05). Expression of GATA transcription factors in mice inhibits differentiation of preadipocytes into mature adipocytes. LPAR1 also inhibits differentiation of preadipocytes in mice, and LPAR1 knock-out mice become significantly fatter than wild-type mice. A SNP that introduces a GATA site in the promoter of LPAR1 could up-regulate its expression in the Lean line, and increased LPA signaling could then inhibit preadipocyte differentiation. Conversely, loss of the GATA binding site could explain decreased levels of LPAR1 expression and attenuated inhibition of adipocyte maturation in the Fat line.