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Title:Identification and characterization of thiamine biosynthesis, transport, and regulation elements among bacteroides species
Author(s):Costliow, Zachary Andrew
Director of Research:Degnan, Patrick H
Doctoral Committee Chair(s):Degnan, Patrick H; Vanderpool, Carin
Doctoral Committee Member(s):Olsen, Gary; Imlay, James
Department / Program:Microbiology
Discipline:Microbiology
Degree Granting Institution:University of Illinois at Urbana-Champaign
Degree:Ph.D.
Genre:Dissertation
Subject(s):Thiamine, gut, microbiome, bacteroides
Abstract:Thiamine (vitamin B1) is an essential cofactor for all organisms. Humans primarily acquire thiamine through their diet and thiamine deficiencies have adverse neurological effects. However, the role gut microbes play in modulating thiamine availability is poorly understood. In addition, little is known about how thiamine, the Bacteroidetes ability to biosynthesize and transport thiamine, or the regulation of biosynthesis and transport impacts the stability of microbial gut communities and human health as a whole. To investigate the role thiamine plays in the gut we leveraged in silico analyses of gut microbial species to determine prominent strategies utilized to attain thiamine. In addition, we have identified the genetic content and operon structure of thiamine transport and biosynthesis across the prominent gut phylum, Bacteroidetes. Along with the bioinformatic methods, RNAseq revealed differential responses to exogenous thiamine by three abundant Bacteroides species. This is highlighted by the global down-regulation of thiamine and amino acid biosynthesis, central, and purine metabolism when thiamine was present in Bacteroides thetaiotaomicron. In contrast Bacteroides uniformis and vulgatus show a much more reserved transcriptomic response to exogenous thiamine. In order to build upon these data, we leveraged genetic mutants of thiamine biosynthesis and transport loci in B. thetaiotaomicron. These analyses determined both systems were critical for growth in thiamine-deficient medium. The defect in the double transport mutant suggests an uncharacterized feedback mechanism between thiamine transport and biosynthesis in B. thetaiotaomicron. Along with the phenotypic analysis of thiamine acquisition operons in B. thetaiotaomicron we investigated how these deletions impacted its fitness in thiamine deplete and replete conditions. Building on the phenotypic analysis in B. thetaiotaomicron we turned our attention to the conserved regulatory TPP riboswitches preceding thiamine biosynthesis and transport genes in B. thetaiotaomicron, B. uniformis, and B. vulgatus utilizing transcriptional and translational reporter assays. These assays have shown a clear regulatory hierarchy between thiamine biosynthesis and transport pathways. In addition, trends in TPP riboswitch distance from their predicted regulon points to the mechanism of regulation (transcriptional or translational) and warrants further investigation. Together, these data show that thiamine acquisition mechanisms and their regulation are critical to physiology and fitness among the Bacteroidetes. In addition future work may provide insight into modeling how other gut microbes respond to the shifting availability of thiamine in the gut and how to therapeutically alter the gut microbiota from a dysbiotic state to a non-disease state.
Issue Date:2018-07-27
Type:Thesis
URI:http://hdl.handle.net/2142/102879
Rights Information:Copyright 2018 Zachary Costliow
Date Available in IDEALS:2019-02-08
Date Deposited:2018-12


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