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QseA regulation of virulence factors in EHEC

ABSTRACT: Enterohemorrhagic E. coli (EHEC) colonizes the large intestine and causes attaching and effacing lesions (AE). Most of the genes involved in the formation of AE lesions are encoded within a chromosomal pathogenicity island termed the Locus of Enterocyte Effacement (LEE). The LysR-like transcriptional factor QseA regulates the LEE by binding directly to the regulatory region of ler. Here, we performed transcriptome analyses comparing WT EHEC and the isogenic qseA mutant in order to elucidate the extent of QseA’s role in gene regulation in EHEC. The following results compare genes that were up-regulated and down-regulated ! 2-fold in the qseA mutant strain compared to the WT strain. At mid-exponential growth, 222 genes were up-regulated and 1874 were downregulated. At late-exponential growth, a total of 55 genes were up-regulated and 605 genes were down-regulated. During mid-exponential growth, QseA represses its own transcription, whereas during late-logarithmic growth, QseA activates expression of the LEE genes as well as non-LEE encoded effector proteins. During both growth phases, several genes carried in O-islands, were activated by QseA, whereas genes involved in cell metabolism were repressed. We also performed electrophoretic mobility shift assays, competition experiments, and DNAseI footprints, and the results suggested that QseA directly binds both the ler proximal and distal promoters, its own promoter, as well as promoters of genes encoded in EHEC-specific O-islands. Additionally, we mapped the transcriptional start site of qseA, leading to the identification of two promoter sequences. Taken together, these results indicate that QseA acts as a global regulator in EHEC, coordinating expression of virulence genes.

ORGANISM(S): Escherichia coli

PROVIDER: GSE18118 | GEO | 2010/09/07

SECONDARY ACCESSION(S): PRJNA119441

REPOSITORIES: GEO

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Project description:RpoS is a conserved stress regulator that plays a critical role in survival under stress conditions in Escherichia coli and other γ-proteobacteria. RpoS is also involved in virulence of many pathogens including Salmonella and Vibrio species. Though well characterized in non-pathogenic E. coli K12 strains, the effect of RpoS on transcriptome expression has not been examined in pathogenic isolates. E. coli O157:H7 is a serious human enteropathogen, possessing a genome 20% larger than that of E. coli K12, and many of the additional genes are required for virulence. The genomic difference may result in substantial changes in RpoS-regulated gene expression. To test this, we compared the transcriptional profile of wild type and rpoS mutants of the E. coli O157:H7 EDL933 type strain. The rpoS mutation had a pronounced effect on gene expression in stationary phase, and more than 1,000 genes were differentially expressed (two-fold, p<0.05). By contrast, we found 11 genes expressed differently in exponential phase. Western blot analysis revealed that, as expected, RpoS level was low in exponential phase and substantially increased in stationary phase. The defect in rpoS resulted in impaired expression of genes responsible for stress response (e.g., gadA, katE and osmY), arginine degradation (astCADBE), putrescine degradation (puuABCD), fatty acid oxidation (fadBA and fadE), and virulence (ler, espI and cesF). For EDL933-specific genes on O-islands, we found 50 genes expressed higher in wild type EDL933 and 49 genes expressed higher in the rpoS mutants. The protein levels of Tir and EspA, two LEE-encoded virulence factors, were elevated in the rpoS mutants under LEE induction conditions. Our results show that RpoS has a profound effect on global gene expression in the pathogenic strain O157:H7 EDL933, and the identified RpoS regulon, including many EDL933-specific genes, differs substantially from that of laboratory K12 strains. In this study, we characterized the RpoS regulon of E. coli O157:H7 strain EDL933 using microarray analysis.

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QseA regulation of virulence factors in EHEC

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