Animals evolved in a world pre-dominated by microscopic organisms. Colonization of intestinal tracts at birth by microbes initiates the next generation of an ancient symbiosis that profoundly impacts our physiology and pathophysiology. A record of this symbiosis is encoded in our genomes. In this dissertation, I explore how regulatory regions embedded in non-genic DNA mediate transcriptional responses to the intestinal microbiota. Extensive research has demonstrated that the complex community of microbes residing within our intestine (the gut microbiota) contributes biochemistries that enhance nutrient digestion, metabolize xenobiotics, and collectively function as an important environmental factor that modulates host energy balance and immunity. However, the mechanisms that host cells use to perceive and respond to these microbial activities are not well understood. I used the zebrafish and mouse gnotobiotic models to define mechanisms by which the microbiota regulates host transcription in the intestinal epithelium at the single gene and genome-wide scales. The intestinal microbiota enhances dietary energy harvest leading to increased lipid storage in peripheral tissues. This effect is caused in part by the microbial suppression of intestinal expression of a circulating inhibitor of Lipoprotein lipase called Angiopoietin-like 4 (Angptl4/Fiaf). I utilized the zebrafish in which host regulatory DNA can be rapidly analyzed in a live, transparent, and gnotobiotic vertebrate to define the cis-regulatory mechanisms controlling angptl4 transcription. I discovered an intronic cis-regulatory module (CRM) that confers intestine-specific transcription and microbial suppression of angptl4. I used comparative sequence analysis from 12 fish species, functional mapping, and mutagenesis to define the minimal set of regulatory sequences required for activity of the angptl4 intestinal CRM. I applied computational prediction and DNA affinity chromatography to discern candidate transcription factors regulating angptl4 intestinal expression. At the genomic level, I employed DNase-seq and FAIRE-seq in the intestine of germ-free and conventionally-raised mice and zebrafish to facilitate the discovery of CRMs meditating host responses to the microbiota genome-wide. This work provides a novel paradigm for understanding how microbial signals interact with tissue-specific regulatory networks to control host gene expression and elucidates mechanisms mediating over 500 million years of co-existence and co-evolution of vertebrate hosts with their intestinal microbiota.
|Advisor:||Rawls, John F.|
|Commitee:||Bultman, Scott J., Lieb, Jason D., Lund, Pauline K., Marzluff, William F., Sethupathy, Praveen|
|School:||The University of North Carolina at Chapel Hill|
|Department:||Genetics & Molecular Biology|
|School Location:||United States -- North Carolina|
|Source:||DAI-B 74/05(E), Dissertation Abstracts International|
|Subjects:||Molecular biology, Genetics, Evolution and Development|
|Keywords:||Angiopoietin, Cis-regulatory module, Fasting induced adipose factors, Intestinal epithelium, Microbiota, Transcription factors|
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