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Molecular Mechanisms Mediating the Effects of Acute Dietary Vitamin A Washington University in St. Louis Washington University Open Scholarship Arts & Sciences Electronic Theses and Dissertations Arts & Sciences Summer 8-15-2016 Molecular Mechanisms Mediating the Effects of Acute Dietary Vitamin A Deficiency on a Defined Model Human Gut Microbiota Matthew hC arles Hibberd Washington University in St. Louis Follow this and additional works at: https://openscholarship.wustl.edu/art_sci_etds Recommended Citation Hibberd, Matthew Charles, "Molecular Mechanisms Mediating the Effects of Acute Dietary Vitamin A Deficiency on a Defined Model Human Gut Microbiota" (2016). Arts & Sciences Electronic Theses and Dissertations. 854. https://openscholarship.wustl.edu/art_sci_etds/854 This Dissertation is brought to you for free and open access by the Arts & Sciences at Washington University Open Scholarship. It has been accepted for inclusion in Arts & Sciences Electronic Theses and Dissertations by an authorized administrator of Washington University Open Scholarship. For more information, please contact [email protected]. WASHINGTON UNIVERSITY IN ST. LOUIS Division of Biology and Biomedical Sciences Molecular Microbiology and Microbial Pathogenesis Dissertation Examination Committee: Jeffrey I. Gordon, Chair Gautam Dantas Daniel E. Goldberg Jeffrey P. Henderson Scott J. Hultgren Lora L. Iannotti Clay F. Semenkovich Molecular Mechanisms Mediating the Effects of Acute Dietary Vitamin A Deficiency on a Defined Model Human Gut Microbiota by Matthew Charles Hibberd A dissertation presented to the Graduate School of Arts and Sciences of Washington University in partial fulfillment of the requirements for the degree of Doctor of Philosophy August 2016 St. Louis, Missouri © 2016, Matthew Charles Hibberd Table of Contents Acknowledgments .......................................................................................................................... iv Abstract of the Dissertation .......................................................................................................... vii Chapter 1 Introduction The microbiota and micronutrient deficiencies in the context of global health ...............................2 Iron deficiency .....................................................................................................................3 Zinc deficiency .....................................................................................................................4 Vitamin A deficiency ............................................................................................................4 Folate deficiency ..................................................................................................................5 Multiple micronutrient deficiencies .....................................................................................5 Studies of micronutrient deficiency in gnotobiotic animal models .................................................6 B vitamins ............................................................................................................................7 Vitamins K and A .................................................................................................................8 Minerals ...............................................................................................................................8 Controlled manipulation of interactions between diet, the gut microbiota, and host: gnotobiotics, defined communities, and assay methods ........................................................................................9 Overview of the dissertation and hypotheses .................................................................................11 References ......................................................................................................................................13 Chapter 2 Molecular mechanisms mediating the effects of acute dietary vitamin A deficiency on a model human gut microbiota Abstract ..........................................................................................................................................24 Introduction ....................................................................................................................................25 Results ............................................................................................................................................27 Dietary vitamin A deficiency has the greatest effect on community structure and meta- transcriptome ......................................................................................................................28 Identification of a member of the TetR family of transcriptional repressors that mediates the retinol sensitivity of B. vulgatus ................................................................................29 ii Characterizing the regulon controlled by AcrR ................................................................32 Further evidence that the AcrAB-TolC efflux system affects retinol sensitivity and the effects of bile acids ...........................................................................................................35 Prospectus ..........................................................................................................................37 Acknowledgements ........................................................................................................................38 Materials and Methods ...................................................................................................................39 References ......................................................................................................................................54 Figure Legends ...............................................................................................................................60 Figures............................................................................................................................................63 Supplemental Materials .................................................................................................................70 Supplemental Figure Legends ........................................................................................................70 Supplemental Figures .....................................................................................................................71 Supplemental Tables ......................................................................................................................75 Chapter 3 Future directions Introduction ....................................................................................................................................82 Further mechanistic characterization of bacterial responses to micronutrients ............................82 Vitamin A resistance mechanisms, toxicity, and interspecies variation in sensitivity .......82 Expanding host-side exploration of bacteria-vitamin A interactions .................................85 The effects of other micronutrient deficiencies on the model gut microbiota ...................87 Technical considerations ���������������������������������������������������������������������������������������������������������������88 Community structure analyses - COPRO-Seq ...................................................................88 Community metatranscriptome analyses - Microbial RNA-Seq .......................................89 Linking gnotobiotic micronutrient deficiency models to real-world global health outcomes .......90 References ......................................................................................................................................93 Figure Legend ................................................................................................................................97 Figure .............................................................................................................................................98 iii Acknowledgments We, though individuals, are products of the collective influences of our environment, and I count myself fortunate to be a product of a number of incredibly supportive and inspiring environments over the course of my personal and professional life. I am proud to count myself a product of the Gordon Lab and the steadfast and enthusiastic guidance of my thesis advisor Jeffrey Gordon. In reflecting on my time in the lab, I am awed by Jeff’s ability to maintain in the lab an ecosystem populated with individuals who possess the professional qualities of creativity, intelligence, and enthusiasm and the personal qualities of empathy, graciousness, and generosity. Jeff leads these traits truly by example and expects that we hold ourselves to the same standard. In return, we are rewarded by an environment full of opportunity and support at a scale it takes (at least) the length of a PhD to truly comprehend. I feel incredibly fortunate to have had the opportunity to train under Jeff’s guidance and learn alongside and from the extraordinarily talented individuals with whom I have shared the last many years in the lab. I would also like to thank my thesis committee, in particular Daniel Goldberg, who de- serves special gratitude for serving as my committee chair. I will always be grateful to my entire committee for their patient guidance and for the time each member devoted to attending my com- mittee meetings and advising my work. The opportunity to learn from a number
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