Location: Produce Safety and Microbiology Research
Title: FdhTU-modulated formate dehydrogenase expression and electron donor availability enhance recovery of Campylobacter jejuni following host cell infection Authors
Submitted to: Journal of Bacteriology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: May 17, 2012
Publication Date: July 18, 2012
Repository URL: http://jb.asm.org/content/194/15/3803.abstract
Citation: Pryjma, M., Apel, D., Parker, C., Huynh, S., Gaynor, E.C. 2012. FdhTU-modulated formate dehydrogenase expression and electron donor availability enhance recovery of Campylobacter jejuni following host cell infection. Journal of Bacteriology. 194:3803-3813. 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 fdhT (cjj81176_1492) and fdhU (cjj81176_1493). This operon consists of a putative membrane transporter (fdhT) and a likely DNA binding protein (fdhU). To explore these genes further, targeted non-polar insertional mutants delta fdhT or delta fdhU were generated and tested for their role in intestinal epithelial cell interactions. In contrast, fluorescence microscopy indicated no defects for delta fdhT or delta fdhU mutants in either adherence or invasion. Each mutant was defective for intracellular survival as assessed by colony forming unit enumeration. Orthology suggests that FdhU 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, delta fdhU, and complemented background were analyzed. The array revealed a ten-fold induction of the formate dehydrogenase operon in a delta fdhU background suggesting FdhU is a repressor of fdhABC transcription. Analysis of a delta fdhA strain revealed a defect in tissue culture invasion similar to delta fdhU suggesting a link between dis-regulation of the fdhABC operon in a delta fdhU background and a lack of formate dehydrogenase activity. Analysis of bacterial growth in the presence or absence of formate in WT, delta fdhU and delta fdhA backgrounds suggest that both delta fdhU and delta 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: Analysis of Campylobacter jejuni fdhTU reveals a role in formate dehydrogenase activity and implications for electron donor requirements during the pathogen-host cell interaction. Campylobacter jejuni is a foodborne bacterial pathogen which colonizes the intestinal tract and causes severe gastroenteritis. Interaction with host epithelial cells is thought to enhance severity of disease, and the ability of C. jejuni to modulate its metabolism in different in vivo and environmental niches contributes to its success as a pathogen. An operon comprising two co-transcribed genes we have designated fdhT (cjj81176_1492) and fdhU (cjj81176_1493) is conserved in many bacterial species. This operon was found to be important for C. jejuni adherence to and/or invasion of Caco-2 epithelial cells as assessed by CFU enumeration. In contrast, fluorescence microscopy indicated no defects for delta fdhT or delta fdhU mutants in either adherence or invasion. Loss of fdhU resulted in lower mRNA levels of several formate dehydrogenase (FDH) genes and a severe defect in FDH activity. Cell infection phenotypes of a mutant deleted for the FdhA subunit of FDH and an delta fdhU delta fdhA double mutant likewise suggested that FdhU and FdhA function in a similar pathway. To reconcile whether CFU recovery defects observed for delta fdhU and delta fdhA mutants represented a defect in intracellular survival or a defect in growth following liberation from the intracellular environment, assays were conducted using recovery plates supplemented with an alternate electron donor, sodium sulfite. Under these conditions, mutant and wildtype strains were recovered equally, and at significantly higher levels than on unsupplemented plates. This work identifies FdhTU and FDH as important for recovery following cell infection and suggests that C. jejuni alters its metabolic potential in the intracellular environment.