Phase variation in <i>tcpH</i> modulates expression of the ToxR regulon in <i>Vibrio cholerae</i>

Carroll, P.; Tashima, Karen T.; Rogers, Marc B.; DiRita, Victor J.; Calderwood, Stephen B.

doi:10.1046/j.1365-2958.1997.5371901.x

Cited by 124 publications

(133 citation statements)

References 40 publications

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“…This conclusion is consistent with the previous report that TcpPH is expressed in a growthdependent manner, i.e., only expressed in mid-logarithmic phase (38). These findings also suggest that the major bile acids are not the component within bile that accounts for the previously described inhibition of CT expression but rather that some other component within the crude mixture accounts for that activity.…”

Section: Resultssupporting

confidence: 92%

Bile acids induce cholera toxin expression in Vibrio cholerae in a ToxT-independent manner

Hung

Mekalanos²

2005

Proc. Natl. Acad. Sci. U.S.A.

149

128

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The production of cholera toxin (CT) during Vibrio cholerae infection results in the hallmark diarrhea that characterizes the disease cholera. The transmembrane protein ToxR was originally identified as a functional transcriptional activator of ctxAB in a heterologous Escherichia coli system. However, direct ToxR activation of the ctxAB promoter in V. cholerae has not been previously demonstrated. Instead, a regulatory cascade has been defined in which the activators ToxRS and TcpPH modulate ctxAB expression by acting in concert to transcriptionally activate another regulator, ToxT. ToxT, in turn, directly activates ctxAB expression as well as expression of the tcp genes and other virulence-associated genes. In this study, we show that ToxRS directly activates ctxAB in a ToxT-independent manner in a classical biotype V. cholerae, and that this activation requires the presence of bile acids. Although the levels of CT induced by this mechanism are lower than levels induced under other in vitro conditions, the bile-dependent conditions described here are more physiologic, being independent of pH and temperature. We further show that the inability of bile acids to stimulate ToxRS-dependent expression of CT in El Tor biotype strains is related to the differences between classical and El Tor ctxAB promoters, which differ in the number of heptad TTTTGAT repeats in their respective upstream regions. The ability of bile acids to stimulate direct activation of ctxAB by ToxRS depends upon the transmembrane domain of ToxR, which may interact with bile acids in the inner membrane of V. cholerae. The Gram-negative bacterium Vibrio cholerae is the infectious agent that causes the severe human diarrheal disease cholera (1). Two major virulence factors, cholera toxin (CT) and the toxin-coregulated pilus (TCP), play major roles in the pathogenesis of this infection. Secretion of CT, an ADP-ribosylating toxin encoded in the genome of the filamentous, lysogenic CTX⌽ phage, results in elevated cAMP levels in intestinal epithelial cells and subsequent secretory diarrhea (2). The regulation of CT and TCP expression has been studied intensely and has been reported to respond to temperature, pH, osmolarity, bile salts, and certain amino acids in vitro (3, 4); nevertheless, a clear understanding of the in vivo environmental stimuli that trigger CT and TCP production remains elusive.The first transcriptional regulator of CT production identified was ToxR, a 32-kDa inner-membrane protein that activates the ctx promoter in Escherichia coli and whose deletion in V. cholerae results in the inability to produce CT (5). Deletion analysis of the ctxAB promoter localized the binding site of ToxR to multiple heptad repeats, TTTTGAT, upstream of ctxAB (6). However, ToxR activation of ctxAB transcription in V. cholerae has not been demonstrated (7). Instead, ToxR in V. cholerae binds with its downstream enhancer ToxS to the upstream region of the toxT gene, which encodes another transcriptional activator (8). ToxT belongs to the AraC family of trans...

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Section: Resultssupporting

confidence: 92%

Bile acids induce cholera toxin expression in Vibrio cholerae in a ToxT-independent manner

Hung

Mekalanos²

2005

Proc. Natl. Acad. Sci. U.S.A.

149

128

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“…AphA and AphB, the latter being a LysR family regulator, were identified as activators of TcpP (Kovacikova and Skorupski, 1999). They couple pH and temperature signals to the expression of CT and TCP via TcpP and ToxT (Carroll et al, 1997); cAMP-CRP couples carbon source to CT and TCP expression by binding to the promoter of tcpP and interfering with AphABdependent activation (Kovacikova and Skorupski, 2001). Much less is known about the combinatorial control of the ToxT-independent branch of the ToxR regulon, except that ToxR may act as a sensor for osmolarity (Miller and Mekalanos, 1988).…”

Section: Co-ordinated Regulation Of the Two Branches Of The Toxr Regumentioning

confidence: 99%

ToxR interferes with CRP‐dependent transcriptional activation of ompT in Vibrio cholerae

Merrell

Camilli

et al. 2002

Molecular Microbiology

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SummaryIn pathogenic Vibrio cholerae, the transmembrane DNA-binding protein ToxR co-ordinates the expression of over 20 genes, including those encoding important virulence factors such as cholera toxin and the toxin-co-regulated pilus. The outer membrane protein OmpT is the only member of the ToxR regulon known to be repressed by ToxR. In this study, we examined the environmental conditions that regulate OmpT expression and demonstrated that ompT transcription is upregulated 14-fold when the bacteria enter late log phase from early log phase. Deletion of the crp gene completely abolishes OmpT expression. Comparison of ompT transcription levels in the isogenic crp -, toxR -and crp -toxR -mutants revealed that (i) in the absence of ToxR, constitutive high-level ompT transcription is dependent on cAMP receptor protein (CRP); (ii) ToxR not only interferes with CRPdependent ompT activation, but also abolishes the CRP-independent, basal level ompT transcription; thus, the mechanism by which ToxR represses ompT transcription involves both antiactivation and direct repression; (iii) both CRP and ToxR are required for the regulation of OmpT expression by growth phase. To provide further insights into the molecular mechanism of CRP-dependent activation of ompT transcription, we demonstrated that CRP-dependent activation requires a CRP binding site centred at -310 of the ompT promoter, without which the interaction of CRP with other CRP binding site(s) more proximal to the promoter results in repression. Mutations in two regions on CRP (AR1 and AR2) that directly

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“…We and others demonstrated that deletion of tcpH from V. cholerae results in poor toxT activation and, consequently, drastic diminution of ctxAB and tcp gene expression (10,16).…”

mentioning

confidence: 94%

“…TcpP interacts with TcpH, which is also a membrane protein but lacks an extensive cytoplasmic domain (9,10). The association of TcpP and TcpH in V. cholerae stabilizes TcpP from degradation, thereby enabling formation of a proposed transcription activation complex of TcpP͞TcpH and ToxR͞ToxS (11)(12)(13).…”

mentioning

confidence: 99%

Degradation of the membrane-localized virulence activator TcpP by the YaeL protease in Vibrio cholerae

Matson

DiRita

2005

Proc. Natl. Acad. Sci. U.S.A.

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132

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A common mechanism inhibiting the activity of transcription factors is their sequestration to the membrane until they are needed, at which point they are released from the membrane by proteolysis. Acting in contrast to this inhibition mechanism are virulence regulators of Vibrio cholerae, the ToxR and TcpP proteins, which are localized to the inner membrane of the cell, where they bind promoter DNA and activate gene expression. TcpP is rapidly degraded in the absence of another protein, TcpH. We used a genetic screen to identify regulators of TcpP stability and identified the YaeL membrane-localized zinc metalloprotease as responsible for degrading TcpP in the absence of TcpH. In Escherichia coli, DegS and YaeL cooperate to degrade RseA, an antisigma factor that sequesters E to the inner membrane, thereby inhibiting the activity of E . When yaeL was disrupted in a V. cholerae tcpH mutant, we observed accumulation of a lower molecular weight species of TcpP. This observation is consistent with TcpP being partially degraded in the absence of YaeL. A mutant lacking both DegS and YaeL continued to accumulate the TcpP degradation product, indicating that protease other than DegS is acting before YaeL in degrading TcpP. The YaeL-dependent degradation pathway is active in TcpH ؉ cells under conditions that are not favorable for virulence gene activation. This work expands the knowledge of YaeL-dependent processing in the bacterial cell and reveals an unexpected layer of virulence gene regulation in V. cholerae.gene regulation ͉ pathogenesis ͉ membrane regulators

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Phase variation in tcpH modulates expression of the ToxR regulon in Vibrio cholerae

Cited by 124 publications

References 40 publications

Bile acids induce cholera toxin expression in Vibrio cholerae in a ToxT-independent manner

Bile acids induce cholera toxin expression in Vibrio cholerae in a ToxT-independent manner

ToxR interferes with CRP‐dependent transcriptional activation of ompT in Vibrio cholerae

Degradation of the membrane-localized virulence activator TcpP by the YaeL protease in Vibrio cholerae

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