Site of transcriptional activation of virB on the large plasmid of Shigella flexneri 2a by VirF, a member of the AraC family of transcriptional activators

Bh, Jost; Adler, Ben

doi:10.1006/mpat.1993.1047

Cited by 23 publications

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“…We have recently reported that pHN280, the pINV plasmid of EIEC HN280, is able to integrate into the host chromosome and that pINV-integrated strains are noninvasive (48). In order to understand the mechanism by which pINVintegrated strains modulate expression of virulence genes, we performed Northern blot analysis to evaluate the transcription of the regulatory genes virB, virF, and hns (virR) (20,23,41,42,46). Equivalent amounts of total RNA extracted from strains harboring an autonomously replicating pINV and from pINVintegrated strains were denatured, separated on 1.2% agarose gels, and hybridized with the virB-, virF-, and hns-specific probes (see Materials and Methods).…”

Section: Resultsmentioning

confidence: 99%

“…Plasmid genes associated with virulence comprise the ipa, mxi, and spa operons that are necessary for invasioin of epithelial cells; the icsA (virG) gene that is essential for intra-and intercellular spread; and the regulatory genes virB and virF (2,16,17,24,46). virB activates transcription of ipa, mxi, and spa and is in turn regulated by two DNA-binding regulatory proteins, VirF and H-NS (1,10,11,16,20,23,26,41,42,46). Mutations or deletions of hns derepress transcription of invasion genes at 30ЊC only in the presence of VirF (10,11,20,23,42).…”

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“…virB activates transcription of ipa, mxi, and spa and is in turn regulated by two DNA-binding regulatory proteins, VirF and H-NS (1,10,11,16,20,23,26,41,42,46). Mutations or deletions of hns derepress transcription of invasion genes at 30ЊC only in the presence of VirF (10,11,20,23,42). VirF positively regulates transcription of virB and icsA in a DNAtopology-dependent manner, while H-NS acts as a negative regulator on both genes (10,42).…”

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H-NS regulation of virulence gene expression in enteroinvasive Escherichia coli harboring the virulence plasmid integrated into the host chromosome

et al. 1995

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We have previously shown that integration of the virulence plasmid pINV into the chromosome of enteroinvasive Escherichia coli and of Shigella flexneri makes these strains noninvasive (C. Zagaglia, M. Casalino, B. Colonna, C. Conti, A. Calconi, and M. Nicoletti, Infect. Immun. 59:792-799, 1991). In this work, we have studied the transcription of the virulence regulatory genes virB, virF, and hns (virR) in wild-type enteroinvasive E. coli HN280 and in its pINV-integrated derivative HN280/32. While transcription of virF and of hns is not affected by pINV integration, transcription of virB is severely reduced even if integration does not occur within the virB locus. This indicates that VirF cannot activate virB transcription when pINV is integrated, and this lack of expression accounts for the noninvasive phenotype of HN280/32. Virulence gene expression in strains HN280 and HN280/32, as well as in derivatives harboring a mxiC::lacZ operon fusion either on the autonomously replicating pINV or on the integrated pINV, was studied. The effect of the introduction of plasmids carrying virB (pBN1) or virF (pHW745 and pMYSH6504), and of a ⌬hns deletion, in the different strains was evaluated by measuring ␤-galactosidase activity, virB transcription, and virB-regulated virulence phenotypes like synthesis of Ipa proteins, contactmediated hemolysis, and capacity to invade HeLa cells. The introduction of pBN1 or of the ⌬hns deletion in pINV-integrated strains induces temperature-regulated expression or temperature-independent expression, respectively, of ␤-galactosidase activity and of all virulence phenotypes, while an increase in virF gene dosage does not, in spite of a high-level induction of virB transcription. Moreover, a wild-type hns gene placed in trans fully reversed the induction of ␤-galactosidase activity due to the ⌬hns deletion. These results indicate that virB transcription is negatively regulated by H-NS both at 30 and at 37؇C in pINV-integrated strains and that there is also a dosedependent effect of VirF on virB transcription. The negative effect of H-NS on virB transcription at the permissive temperature of 37؇C could be due to changes in the DNA topology occurring upon pINV integration that favor more stable binding of H-NS to the virB promoter DNA region. At 30؇C, the introduction of the high-copy-number plasmid pMYSH6504 (but not of the low-copy-number pHW745) or of the ⌬hns deletion induces, in strains harboring an autonomously replicating pINV, ␤-galactosidase activity, virB transcription, and expression of the virulence phenotypes, indicating that, as for HN280/32, the increase in virF gene dosage overcomes the negative regulatory effect of H-NS on virB transcription. Moreover, we have found that virF transcription is finely modulated by temperature and, with E. coli K-12 strains containing a virF-lacZ gene fusion, by H-NS. This leads us to speculate that, in enteroinvasive bacteria, the level of VirF inside the cell controls the temperature-regulated expression of invasion genes.Shigella flexneri and ...

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H-NS regulation of virulence gene expression in enteroinvasive Escherichia coli harboring the virulence plasmid integrated into the host chromosome

et al. 1995

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“…Salmonella Enteritidis contains a potent restriction system that limits genetic manipulations in this strain. (Caron et al, 1989 ;Edwards & Schifferli, 1997 ;Jost & Adler, 1993 ;Nataro et al, 1994 ;Tobe et al, 1996).…”

Section: Transduction Of the Sef Island To Salmonella Typhimurium Lt2mentioning

confidence: 99%

Genomic analysis and growth-phase-dependent regulation of the SEF14 fimbriae of Salmonella enterica serovar Enteritidis The GenBank accession number for the sequence reported in this paper is AF239978.

et al. 2001

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Salmonella enterica serovar Enteritidis is a leading cause of food poisoning in the USA and Europe. Although Salmonella serovars share many fimbrial operons, a few fimbriae are limited to specific Samonella serovars. SEF14 fimbriae are restricted to group D Salmonella and the genes encoding this virulence factor were acquired relatively recently. Genomic, genetic and gene expression studies have been integrated to investigate the ancestry, regulation and expression of the sef genes. Genomic comparisons of the Salmonella serovars sequenced revealed that the sef operon is inserted in leuX in Salmonella Enteritidis, Salmonella Paratyphi and Salmonella Typhi, and revealed the presence of a previously unidentified 25 kb pathogenicity island in Salmonella Typhimurium at this location. Salmonella Enteritidis contains a region of homology between the Salmonella virulence plasmid and the chromosome downstream of the sef operon. The sef operon itself consists of four co-transcribed genes, sefABCD, and adjacent to sefD there is an AraC-like transcriptional activator that is required for expression of the sef genes. Expression of the sef genes was optimal during growth in late exponential phase and was repressed during stationary phase. The regulation was coordinated by the RpoS sigma factor.

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“…The transcription of genes of the entry region from the divergent icsB and ipgD promoters is under the control of both VirF and VirB. The expression of VirF, a member of the AraC family that is encoded by the virulence plasmid, is induced at 37°C (14) and activates transcription from the virB promoter (23). VirB, a member of the ParB family of partitioning proteins, is required for transcription of the icsB and ipgD promoters (1).…”

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Identification of the cis -Acting Site Involved in Activation of Promoters Regulated by Activity of the Type III Secretion Apparatus in Shigella flexneri

2002

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Bacteria of Shigella spp. use a virulence plasmid-encoded type III secretion (TTS) system to invade the colonic epithelium in humans. The activity of the TTS apparatus is tightly regulated in the wild-type strain and is induced upon contact of bacteria with epithelial cells, whereas it is deregulated, i.e., constitutively active, in some mutants. Under conditions of deregulated secretion, approximately 20 proteins are secreted, including VirA, OspB to OspG, and at least three members of the IpaH family, all of which are encoded by the virulence plasmid. Conditions inducing or deregulating the activity of secretion also induce the transcription of virA and four ipaH genes. The transcription of virA and ipaH9.8 requires both MxiE, a transcriptional activator of the AraC family, and IpgC, the chaperone of IpaB and IpaC, acting as a coactivator. Using reporter plasmids containing lacZ transcriptional fusions, we showed that the ipaH7.8, ipa4.5, ospC1, and ospF promoters are activated under conditions of deregulated secretion and that both MxiE and IpgC are necessary and sufficient for their activation in both Shigella flexneri and Escherichia coli. Promoter mapping and deletion analysis of the ipaH9.8, virA, and ospC1 promoters identified a 17-bp motif, the MxiE box, which overlaps the ؊35 region and is essential for the activation of these promoters. The presence of eight MxiE boxes on the virulence plasmid suggests that 11 genes encoding secreted proteins may be regulated by the activity of secretion. We also present evidence that at least one ipaH gene that is carried by the chromosome is controlled by MxiE and IpgC.

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Site of transcriptional activation of virB on the large plasmid of Shigella flexneri 2a by VirF, a member of the AraC family of transcriptional activators

Cited by 23 publications

References 0 publications

H-NS regulation of virulence gene expression in enteroinvasive Escherichia coli harboring the virulence plasmid integrated into the host chromosome

H-NS regulation of virulence gene expression in enteroinvasive Escherichia coli harboring the virulence plasmid integrated into the host chromosome

Genomic analysis and growth-phase-dependent regulation of the SEF14 fimbriae of Salmonella enterica serovar Enteritidis The GenBank accession number for the sequence reported in this paper is AF239978.

Identification of the cis -Acting Site Involved in Activation of Promoters Regulated by Activity of the Type III Secretion Apparatus in Shigella flexneri

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