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United States Department of Agriculture

Agricultural Research Service

Research Project: Molecular Biology of Human Pathogens Associated with Food

Location: Produce Safety and Microbiology Research

Title: The civRT operon is important for Campylobacter jejuni strain 81-176 host cell interactions through regulation of the formate dehydrogenase operon

Authors
item Pryjma, Mark -
item Parker, Craig
item Apel, Dmitry -
item Gaynor, Erin C -

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: June 1, 2011
Publication Date: N/A

Interpretive Summary: C. jejuni colonizes the mucous layer of the intestine, and the severity of disease by different strains is correlated with host cell interaction and invasion. A microarray-based gene transcription screen to identify genes differentially regulated during C. jejuni interaction with tissue culture cells revealed the up-regulation of a two gene operon termed civRT (Campylobacter Intercellular Viability Regulator/Transporter). This operon consists of a putative membrane transporter (civT) and a likely DNA binding protein (civR). To explore these genes further, targeted non-polar insertional mutants 'civT and 'civR were generated and tested for their role in intestinal epithelial cell interactions. Each mutant was defective for intracellular survival. Orthology suggests that CivR may act as a regulator, and this operon was already observed to exhibit expression changes during infection. To explore the regulon further, a microarray comparing relative transcription levels in WT, 'civR, and complemented background were analyzed. The array revealed a ten-fold induction of the formate dehydrogenase operon in a 'civR background suggesting CivR is a repressor of fdhABC transcription. Analysis of a 'fdhA strain revealed a defect in tissue culture invasion similar to 'civR suggesting a link between dis-regulation of the fdhABC operon in a 'civR background and a lack of formate dehydrogenase activity. Analysis of bacterial growth in the presence or absence of formate in WT, 'civR and 'fdhA backgrounds suggest that both 'civR and 'fdhA lack the ability to use formate as an electron donor, further supporting this hypothesis. Previous work has shown that changes in gene regulation occur upon C. jejuni interaction with host cells. Identifying and understanding regulatory units controlling the transition to an infective state will yield a greater understanding of how C. jejuni causes disease.

Technical Abstract: C. jejuni colonizes the intestinal mucosa, and the severity of disease in different strains is correlated with host cell interaction and invasion. A microarray screen to identify genes differentially regulated during C. jejuni interaction with tissue culture cells revealed the up-regulation of a two gene operon termed civRT (Campylobacter Intercellular Viability Regulator/Transporter). This operon consists of a putative membrane transporter (civT) and a likely DNA binding protein (civR). To explore these genes further, targeted non-polar insertional mutants 'civT and 'civR were generated. To test the role of these genes in intestinal epithelial cell interactions, 'civT and 'civR were used to infect Caco-2 tissue culture cells in a gentamycin protection assay. Each mutant was defective for intracellular survival. Orthology suggests that CivR may act as a regulator, and this operon was already observed to exhibit expression changes during infection. To explore the regulon further, a microarray comparing relative transcription levels in WT, 'civR, and complemented background were analyzed. The array revealed a ten-fold induction of the formate dehydrogenase operon in a 'civR background suggesting CivR is a repressor of fdhABC transcription. Analysis of a 'fdhA strain revealed a defect in tissue culture invasion similar to 'civR suggesting a link between dis-regulation of the fdhABC operon in a 'civR background and a lack of formate dehydrogenase activity. Analysis of bacterial growth in the presence or absence of formate in WT, 'civR and 'fdhA backgrounds suggest that both 'civR and 'fdhA lack the ability to use formate as an electron donor, further supporting this hypothesis. Previous work has shown that changes in gene regulation occur upon C. jejuni interaction with host cells. Identifying and understanding regulatory units controlling the transition to an infective state will yield a greater understanding of how C. jejuni causes disease.

Last Modified: 4/15/2014
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