Regulation of the vibrio cholerae msha pilus drives host colonization and pathogenesis

This federal Project Grant award from the National Institute of Allergy and Infectious Diseases (NIAID) under the Allergy and Infectious Diseases Research program (CFDA 93.855) provides $822,209 to Illinois State University to conduct research on the regulation of the Vibrio cholerae MSHA pilus and its role in host colonization and pathogenesis.

The key objectives of this 3-year research project are to: 1) characterize the promoter regions of the MSHA operons and evaluate the regulation of MSHA pilus production by the FadR fatty acid metabolism regulator, and 2) assess the impact of host-derived long-chain fatty acids on FadR activity and MSHA pilus expression, surface colonization, and biofilm formation. This research aims to elucidate novel host-responsive regulatory mechanisms that facilitate Vibrio cholerae's transitions between environmental and host niches, which could inform strategies for attenuating the pathogen's environmental persistence and host infectivity.

Abstract

Development of multicellular biofilm communities is vital to the lifecycle of many bacterial species. Biofilm formation begins with the attachment of a motile bacterium to a surface, though the underling mechanisms that regulate surface attachment in most biofilm-forming bacterial species remain to be elucidated. For the pandemic cholera pathogen vibrio cholerae, proper regulation of the type IV mannose-sensitive hemagglutinin (msha), is a key factor for mediating the transition from environmental colonization to host infection and pathogenesis. Msha pilus production is essential for v. Cholerae environmental surface attachment and biofilm formation, but cell-surface exposure of msha pili in the host results in bacterial clearance, preventing host infection and pathogenesis. In v. Cholerae, msha pilus components are projected to be encoded within three distinct genetic operons; an assembly operon (MSH-i), a structural operon (MSH-II), and a retraction operon (piltu). However, little is known about how these MSH operons are expressed or regulated. Given that proper regulation of msha pilus expression/production is vital for not only environmental survival, but for host colonization and infection, the goal of my research group is to define mechanisms and regulatory pathways that mediated msha pilus production to facilitate these vital niche transitions during the v. Cholerae lifecycle. An initial screen for regulators of msha production, identified the fatty acid metabolism regulator protein fadr as a putative positive regulator (enhancer) of MSH-i operon expression under environmental conditions. Enhancer activity of fadr is negatively- controlled by exogenously-derived long-chain fatty acids (lcfas), which are a large component of the mammalian diet, and can readily be taken up from the environment by v. Cholerae. Our preliminary data demonstrates that both deletion of fadr and supplementation of known fadr lcfa ligands, significantly diminishes MSH-i operon expression. Our working hypothesis is that fadr promotes expression of the MSH-i operon to facilitate msha pilus production and surface colonization under environmental conditions, and that host-derived lcfas suppress fadr enhancer activity to down-regulate msha production and promote v. Cholerae host colonization and infection. This proposal seeks to address this hypothesis, through two specific aims. In aim 1, we will characterize the promoter regions of the MSH operons, and evaluate fadr-mediated regulation of MSH-i operon expression and msha pilus production. We will then seek to assess the role of fadr lcfa ligands on the regulation of fadr enhancer activity of MSH-i expression, msha pilus production, and v. Cholerae surface colonization and biofilm formation as a part of aim 2. The outcome of this work will elucidate a novel host-responsive regulatory mechanism that facilitates transitions in niche colonization during the v. Cholerae lifecycle, and could lead to the development of strategies for attenuation of v. Cholerae environmental persistence and/or host infectivity.