<p> Pregnane X receptor (PXR) is a major transcriptional regulator of xenobiotic metabolism and transport pathways in the liver and intestines, which are critical for protecting organisms against potentially harmful xenobiotic and endobiotic compounds. Inadvertent activation of drug metabolism pathways through PXR is known to contribute to drug resistance, adverse drug-drug interactions, and drug toxicity in humans. In both humans and rodents, PXR has been implicated in non-alcoholic fatty liver disease, diabetes, obesity, inflammatory bowel disease, and cancer. Because of PXR's important functions, it has been a therapeutic target of interest for a long time.</p><p> Recent mechanistic studies have shown that PXR is modulated by multiple PTMs. In this thesis work, we conducted the first detailed examination of PXR regulation by acetylation. We found that PXR is efficiently acetylated <i> in vitro</i> and <i>in vivo</i> in multiple cell lines (293T, HepG2, LS174T). Acetylation and deacetylation are mediated by p300 and SIRT1, respectively. We found that PXR is directly acetylated by p300 at K109 by LC-MS/MS analysis. The K109Q acetylation mimicking mutant displayed reduced transcriptional activity and reduced ability to induce <i>cyp3A4</i> target gene mRNA and protein compared to the WT and the K109R acetylation-defective mutant. The diminished activity of the K109Q mutant appears to be due to impaired heterodimerization with RXRa and impaired binding of the PXR-RXRa heterodimer to DNA response elements. Furthermore, PXR acetylation appears to have an effect at the phenotypic level, in that pharmacological modulation of PXR acetylation levels can modulate its lipogenic function in mouse primary hepatocytes independent of a ligand. Moreover, the K109Q mutant displays impaired chemoprotective function based on morphological assessment of cells overexpressing K109Q and challenged with indomethacin, suggesting that K109 acetylation downregulates PXR's chemoprotective and perhaps anti-apoptotic functions, although this must be explored further. Notably, the K109R mutant displayed the WT phenotype, further supporting that acetylation itself, not just any arbitrary mutation confers the effect. Altogether, the data suggests that acetylation at K109 represents an overall "loss of function" effect on PXR activity. Implications of our findings are discussed in the context of known roles for PXR in transcription, health, and disease.</p>

<p> The focus of the present research is the synthesis of polypeptides for the study of protein folding and misfolding and for the development of novel polypeptide-based optical antennas in nanotechnology. It is hypothesized that simple polypeptides can be used as models to mimic <i>in vivo</i> folding of globular proteins. Desired repetitive polypeptides were genetically encoded and expressed in <i>E. coli</i> using conventional methods and characterized using a variety of spectroscopic (including circular dichroism (CD), deep UV resonance Raman (DUVRR), UV-vis and fluorescence) and microscopic (atomic force microscopy (AFM) and transmission electron microscopy (TEM)) techniques. The polypeptides predominantly formed bilayer, fibrillar structures with a cross &beta;-core. <b>YEHK21-YE8</b>, a chimeric polypeptide, folded within three days. The folding/fibrillation of the chimeric construct illuminates the controlling factors and hence suggests the importance of those factors in amyloidogenic diseases. <b>YE8</b> and <b>YE8</b> derivatives illustrated the role of proline in &beta;-sheet formation. The EW polypeptide models elucidated the influence of tryptophan residues and the degree of polymerization on folding. The study of <b>EW14C1</b> and <b>EW21C1</b> demonstrated light-harvesting properties when labeled with a suitable dye.</p>

<p> During leaf senescence nutrients are mobilized towards newly developing vegetative and reproductive structures. The IAOx pathway that produces auxin and defense molecules [indole glucosinolates (IGs) and camalexin] is up-regulated during senescence. To investigate the role of the IAOx metabolites we isolated two independent cyp79B2/cyp79B3 double mutants, which are deficient in IGs and camalexin and had reduced auxin levels. Chlorophyll, protein, and gene expression data indicate that cyp79B2/cyp79B3 mutants display early leaf senescence. Furthermore, leaves accumulated higher levels of hydrogen peroxide and seed production was significantly reduced. Auxin signaling at hydathodes and vascular tissue decreased as leaves aged, even though endogenous auxin levels increased. Since CYP79B2/CYP79B3 play only a minor role in auxin synthesis, it is most likely that IGs and/or camalexin are playing a protective role during age-induced developmental leaf senescence. Identifying molecules that slow down the rate of senescence may allow for genetic manipulation to increase nutritional value and crop yield</p>

<p>Soybean (Glycine max) yield is significantly affected by soybean cyst nematode (SCN), Heterodera glycines, and causes an annual loss of billions of US dollars. In this study, Glycine max xyloglucan endotransglycosylase/hydrolase gene (Gm-XTH52) was transformed into a nematode susceptible G. max [Williams 82/PI 518671] variety of soybean to test whether the protein expression has a role in resistance to H. glycines, and possible chemical changes the expression may cause in the plant composition. Expression level of the Gm-XTH52 gene was three times higher than in controls. Significant reduction in the number of SCN cysts suggested suppression of H. glycines parasitism upon transformation. While total sugar amounts did not significantly differ between the transformed and control plants, xyloglucan amounts of loosely bound sugars of genetically mosaic plants were significantly lower in comparison to controls. Control plants showed lower molecular weight sugars than the transformed plants not subjected to H. glycines infection.

<p> Genome sequencing technology has enabled the identification of genetic variants that are linked with cancer phenotypes, whether these are somatically acquired mutations or common inherited single nucleotide polymorphisms (SNPs). Whereas coding variants have been reported to disrupt protein function to promote cancer, most variants map to noncoding regions, with no known function. Recently, much effort has gone into annotating the human noncoding genome, enabling the characterization of the functional basis of noncoding SNPs. As an example of functional impact, breast cancer (BrCa) risk-associated SNPs can alter transcription factor binding at distal enhancers. </p><p> Identifying the targets of risk SNPs remains a challenge. One reason for this is the complex three-dimensional structure of the genome. Local chromatin openness correlates with chromatin activity, and sites of chromatin that are open concurrently across multiple cell types indicates a functional relationship between them. We mapped BrCa risk-associated SNPs to regions of open chromatin to predict the most likely functional risk SNPs. Then, we predicted their targets by identifying the gene promoters whose openness correlated with these risk regions. Further, we validated a gene which is a novel therapeutic target and relevant in breast cancer biology. </p><p> In addition to SNPs, noncoding somatic mutations are also predicted to play a role in cancer. In 2012, driver mutations were reported in the telomerase gene promoter, hinting at the relevance of mutations in regulatory elements. This is particularly true when considering oncogenes whose elevated expression in certain cancers is not attributable to coding mutations or copy number amplification. We reveal the enrichment and functional nature of somatic mutations mapping to enhancers that regulate the estrogen receptor gene, which is known to drive over two-thirds of breast cancer. </p><p> Attributing function to noncoding SNPs and mutations associated with cancer risk and progression is a growing necessity in this era of whole-genome cancer biology. This thesis demonstrates a methodology to identify the functional consequence and gene targets of significantly mutated or risk variant-bearing enhancer sets to narrow the gap between known and unknown risk factors in BrCa.</p>

<p> Breast cancer cell dormancy is a significant clinical problem which contributes to the development of distant metastasis and disease relapse. Currently, no therapies exist which can effectively detect or eradicate dormant cancer cells. </p><p> In this study, we utilized a 3D co-culture dormancy model, recapitulating the inhibitory hematopoietic stem cell niche, which interacts with MDA-MB-231 cells, causing them to enter a state of growth arrest. The knockdown of emerging dormancy regulator gene, p38/MAPK14, in MDA-MB-231 cells allows previously dormant cells to &ldquo;break&rdquo; dormancy and re-enter the cell cycle when grown in the inhibitory niche. Using the newly described in vitro dormancy model, we performed a genomic shRNA library screen, and identified several p38-regulated breast cancer dormancy suppressor gene candidates. Two p38-regulated gene candidates were investigated further. Knockdown of transcription factors and p38 substrates, HBP1 and BHLHB3, in MDA-MB-231 cells lead to re-activation (proliferation) of once indolent cells when cultured in the inhibitory niche. </p><p> The present study illustrates the role of p38 and p38-regulated genes in breast cancer dormancy within the microenvironment of the inhibitory (endosteal) hematopoietic stem cell niche. Additionally, we have identified a list of ~700 breast cancer dormancy suppressor candidate genes. Further analysis and validation experiments are needed to classify novel molecular players and signaling pathways involved in tumor cell dormancy from the list of candidate genes generated in this study.</p>

Thesis (Ph.D.)--Indiana University, Dept. of Biology, 2007. / Source: Dissertation Abstracts International, Volume: 68-05, Section: B, page: 2790. Adviser: Susan Strome. "Title from dissertation home page (viewed Jan. 24, 2008)."

Thesis (Ph.D.)--University of California, San Francisco, 2006. / Source: Dissertation Abstracts International, Volume: 68-01, Section: B, page: 0085. Adviser: Jonathan S. Weissman.

Thesis (Ph.D.)--University of California, San Francisco, 2005. / Source: Dissertation Abstracts International, Volume: 66-12, Section: B, page: 6424. Adviser: Christine Guthrie.

Thesis (Ph.D.)--University of California, San Francisco, 2006. / Source: Dissertation Abstracts International, Volume: 68-01, Section: B, page: 0085. Adviser: Jonathan S. Weissman.