The role of microRNAs, DNA methylation and translational control in regulation of sex specific gene expression in mouse liver

Sex differences are widespread in both mouse and human liver, and are associated with sex differences in drug metabolism and liver pathophysiology. The secretory patterns of growth hormone (GH) is one of the major drivers of liver sex specificity, where intermittent and continuous secretion in male and female respectively lead to sex bias in the expression of more than 1000 genes in mouse liver, via a complex interplay of GH-responsive transcription factors and epigenetic mechanisms. This thesis explores three themes of molecular control in the regulation of liver sex differences: microRNAs, DNA methylation, and translational control. Studies herein identified two microRNAs, miR-1948-5p and miR-802-5p, whose expression is sex biased and regulated by GH and the transcription factor STAT5b. Small RNA sequencing confirmed the sex specificity of these two microRNAs and identified an additional 18 sex-biased microRNAs. Computational and experimental characterization of miR-1948-5p and miR-802-5p confirmed their authenticity. In vivo inhibition of these microRNAs by locked nucleic acids indicated that miR-1948-5p and miR-802-5p played a functional role in repressing female-biased genes and male-biased genes, respectively. This thesis also investigated the impact of GH and STAT5b on liver DNA methylation profiles. Reduced representation bisulfite sequencing was performed on liver tissues from four mouse models that perturbed the GH and STAT5b axis. In the wildtype liver, sex biased demethylation was positively associated with sex biased chromatin opening and gene expression. Global hypermethylation was observed in livers of mice with lit/lit mutation resulting in GH deficiency or with hepatocyte-specific deletion of the STAT5ab locus. Strikingly, these hypermethylated loci were enriched for enhancer elements and STAT5b binding sites found in wild-type mouse liver. Hypophysectomy followed by GH replacement mouse models identified differentially methylated regions that were sex-biased and rapidly methylated and demethylated in response to GH stimulation. Finally, we used ribosome profiling to validate sex-biased protein translation and identify mechanisms of translational control. In sum, this body of work provides novel insights and broadens our understanding of the diverse molecular mechanisms underlying sexual dimorphism in the liver.2020-10-08T00:00:00

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Regulation of drug metabolism and toxicity by multiple factors of genetics, epigenetics, lncRNAs, gut microbiota, and diseases: a meeting report of the 21 st International Symposium on Microsomes and Drug Oxidations (MDO)

Variations in drug metabolism may alter drug efficacy and cause toxicity better understanding of the mechanisms and risks shall help to practice precision medicine. At the 21st International Symposium on Microsomes and Drug Oxidations held in Davis, California, USA, in October 2–6, 2016, a number of speakers reported some new findings and ongoing studies on the regulation mechanisms behind variable drug metabolism and toxicity, and discussed potential implications to personalized medications. A considerably insightful overview was provided on genetic and epigenetic regulation of gene expression involved in drug absorption, distribution, metabolism, and excretion (ADME) and drug response. Altered drug metabolism and disposition as well as molecular mechanisms among diseased and special populations were presented. In addition, the roles of gut microbiota in drug metabolism and toxicology as well as long non-coding RNAs in liver functions and diseases were discussed. These findings may offer new insights into improved understanding of ADME regulatory mechanisms and advance drug metabolism research

Regulation of drug metabolism and toxicity by multiple factors of genetics, epigenetics, lncRNAs, gut microbiota, and diseases: a meeting report of the 21 st International Symposium on Microsomes and Drug Oxidations (MDO)

Variations in drug metabolism may alter drug efficacy and cause toxicity; better understanding of the mechanisms and risks shall help to practice precision medicine. At the 21st International Symposium on Microsomes and Drug Oxidations held in Davis, California, USA, in October 2-6, 2016, a number of speakers reported some new findings and ongoing studies on the regulation mechanisms behind variable drug metabolism and toxicity, and discussed potential implications to personalized medications. A considerably insightful overview was provided on genetic and epigenetic regulation of gene expression involved in drug absorption, distribution, metabolism, and excretion (ADME) and drug response. Altered drug metabolism and disposition as well as molecular mechanisms among diseased and special populations were presented. In addition, the roles of gut microbiota in drug metabolism and toxicology as well as long non-coding RNAs in liver functions and diseases were discussed. These findings may offer new insights into improved understanding of ADME regulatory mechanisms and advance drug metabolism research. (C) 2017 Chinese Pharmaceutical Association and Institute of Materia Medica, Chinese Academy of Medical Sciences. Production and hosting by Elsevier B.V.U.S. National Institutes of Health (NIH) [U01CA175315, R01GM113888]; NIH [ES006694, ES007091, ES021800, ES020522, ES005022, ES023438, DK083952, R01HL122593, R01ES025708, CA098468, T32DK007737, R01DK33765, R01ES024421, R01DK104656, R01DK080440, R01ES025909, R21AA022482, R21AA024935, R01ES019487, R01GM087367, R01GM118367]; Robert Bosch Foundation, Stuttgart, Germany; Searle Scholars Program, USA; VA Merit Award, USA [1I01BX002634]; National Natural Science Foundation of China [81572443]; Yale Liver Center, USA [P30 DK34989]; Amgen; Genentech; Gilead; Pfizer; Bristol-Myers Squibb; UC DavisOpen Access Journal.This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]