All ETDs from UAB

Advisory Committee Chair

Jorge A Benitez

Advisory Committee Members

David E Briles

Kevin Dybvig

Hui Wu

Document Type

Dissertation

Date of Award

2016

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

The diarrheal disease cholera is caused by the Gram-negative and motile bacterium Vibrio cholerae of serogroups O1 and O139. V. cholerae can switch between planktonic (motile) and sessile (biofilm) lifestyles. Biofilms are sessile communities encased in a self-produced extracellular matrix mainly composed of exopolysaccharide, proteins and extracellular DNA. Biofilm formation enhances the capacity of V. cholerae to persist in environmental waters and increases its infectivity. The bacterial second messenger cyclic diguanylic acid (c-di-GMP) regulates the transition between motile and biofilm lifestyles in V. cholerae. At low cell density, two c-di-GMP receptors, the LuxR-type regulator VpsT and the NtrC-type regulator VpsR activate the transcription of vps and rbm genes encoding the exopolysaccharide and protein components of the biofilm matrix, respectively. However, these c-di-GMP signaling events are poorly understood at the level of promoter activation. In this study, we determine for the first time the transcriptome and genome-wide binding profile of the histone-like nucleoid structuring (H-NS) protein in a two-chromosome bacterium. Salient features of the V. cholerae hns transcriptome and binding profile is the clustering of H-NS at chromosomal sites controlling virulence, surface attachment and biofilm development resulting in transcriptional repression. We show that H-NS directly represses the transcription of the vpsA-K and vpsL-Q operons, the rbmABCDEF cluster and vpsT. We demonstrate that in the presence of c-di-GMP, VpsT can disrupt previously formed H-NS nucleoprotein complexes at the vpsA, vpsL and rbm promoters. Artificially increasing the c-di-GMP pool diminished H-NS occupancy at the vpsT promoter and this effect required VpsR. These results support a model in which environmentally-induced fluctuations in the cellular c-di-GMP reversibly control the expression of biofilm-associated genes by triggering a VpsR- and VpsT-dependent H-NS anti-repression cascade. Finally, we note that the clustering of H-NS occupancy along the V. cholerae genome supports a recent view in which gene regulation drives nucleoid architecture. However, we found important differences in the distribution of H-NS clusters between the V. cholerae and E. coli genomes.

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