Comparison of the expression profiles of 630E strain and hfQ mutants
Ontology highlight
ABSTRACT:
ORGANISM(S): Peptoclostridium difficile
SUBMITTER: Monot Marc
PROVIDER: E-GEOD-55274 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Publications
Pleiotropic role of the RNA chaperone protein Hfq in the human pathogen Clostridium difficile.
Boudry P P Gracia C C Monot M M Caillet J J Saujet L L Hajnsdorf E E Dupuy B B Martin-Verstraete I I Soutourina O O
Journal of bacteriology 20140630 18
Clostridium difficile is an emergent human pathogen and the most common cause of nosocomial diarrhea. Our recent data strongly suggest the importance of RNA-based mechanisms for the control of gene expression in C. difficile. In an effort to understand the function of the RNA chaperone protein Hfq, we constructed and characterized an Hfq-depleted strain in C. difficile. Hfq depletion led to a growth defect, morphological changes, an increased sensitivity to stresses, and a better ability to spor ...[more]
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Project description:Clostridium difficile is an emergent human pathogen and the most common cause of nosocomial diarrhea. Our recent deep-sequencing data strongly suggest the importance of RNA-based mechanisms for the control of gene expression in C. difficile. The RNA chaperon protein Hfq is a key component of post-transcriptional regulatory network in various bacterial systems. In an effort to understand the function of Hfq in C. difficile, we have constructed and characterized an Hfq-depleted strain in this bacterium. We present evidence that Hfq is necessary for the normal cell growth, for the establishment of bacillary form and might be involved in metabolic adaptations, stress response and sporulation, important during C. difficile infection cycle. In accordance to these phenotypic changes, our transcriptome analysis revealed pleiotropic effects of Hfq on gene expression in C. difficile. This microarray analysis showed altered expression of genes encoding cell wall and membrane transport components, sporulation proteins, as well as stress response, transcriptional regulator genes and genes of unknown function. Remarkably, a great number of genes belonging to the regulon dependent on sporulation-specific sigma K factor were generally up-regulated in the strain depleted for Hfq. Many important metabolic pathways were also affected by Hfq depletion. Finally, the altered accumulation of several previously identified sRNAs suggests potential involvement of Hfq in these regulatory RNA function. Altogether, these results provide evidence for the pleiotropic role of Hfq protein in C. difficile physiology expanding our knowledge of Hfq-dependent regulation in Gram-positive bacteria. Two-conditions experiments, 630E strain vs. hfQ mutant grown in TYt, 4 biological replicates for each condition.
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Project description:The virulence factors of Clostridium difficile have been studied for many years, but the main in vivo pathogenesis processes of this bacterium has to be investigated. Especially, data on the early mechanisms of C. difficile adaptation to the host is not yet available. The objective of our study was to improve the understanding of the pathogenesis of C. difficile by the analysis of the genome-wide temporal expression of C. difficile genes during the first hours of infection. Three groups of 4 axenic mice each were challenged by vegetative cells of the 630 C. difficile strain, and sacrified at 8, 14, and 38 hours post-infection. Pure prokaryotic RNA was obtained from caecal bacteria. Comparative hybridizations on strain 630 microarrays were done using cDNA issued from an 8-hours in vitro culture as control, with a dye-swap protocol for each sample. Normalisation and statistical analysis of all data were done with different functions of the limma package. The pathogenesis of C. difficile could be seen as a result of the successful metabolic adaptation of the bacteria to its host, contributing to its persistence and multiplication, and the coordinate expression of virulence factors. Analysis of our data enlightened some of these aspects. The results support strongly a two-step infection model, since there is, during the course of infection, a significant increase in the toxins expression contrasting with a decreased expression of most of the putative colonization factors. Several paralogs of the HMW S-layer protein are also down-regulated, some of them could be strong candidates as colonisation factors. Bacterial adaptation to the microenvironment of the host is assessed by the regulation of numerous metabolic pathways, i.e., the upregulation of the ethanolamine catabolic operon or the modulation of several PTS systems. Inactivation of some putative virulence factors identified by this methodology will complete this analysis. Transcriptional profiling of the C. difficile 630E strain after 8h, 14h or 38h in TY. Two-condition experiments: 630E strain 14h or 38h in TY vs. 630E strain 8h in TY. 2 biological replicates for each condition in dye swap.
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