Linked Life Data

11029691

Source:http://linkedlifedata.com/resource/pubmed/id/11029691

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1
pubmed:dateCreated
2000-11-20
pubmed:abstractText
ToxR is required in Vibrio cholerae for transcriptional activation of the toxT gene, the protein product of which activates numerous genes involved in virulence. Although ToxR cannot activate the toxT promoter in Escherichia coli, the products of the tcpPH operon are shown here to activate the toxT promoter, and co-expression with ToxRS enhances activation. An identical pattern was seen in a DeltatcpPDeltatoxR strain of V. cholerae when TcpPH or ToxRS was expressed from plasmids. Although overexpression of the TcpP/H proteins in V. cholerae partially complemented both a DeltatoxR strain and a DeltatcpPDeltatoxR double mutant for toxin production and toxT-lacZ activation, the presence of ToxR greatly increased their expression. Analysis of a toxT-lacZ promoter deletion series demonstrated that TcpP was able to interact functionally with the toxT promoter downstream of the ToxR binding site. This was confirmed using electrophoretic mobility shift assays of this toxT promoter deletion series and DNase I footprinting analysis, which showed that TcpP interacts with the promoter region from -51 to -32, whereas ToxR protected a region from -100 to -69. In addition, membranes containing endogenous levels of ToxR bound more readily to the toxT promoter than did membranes containing only TcpP. Characterization of a number of tcpP substitution mutants revealed one derivative (TcpP-H93L) that, when overexpressed, was markedly defective for toxT activation, cholera toxin and TcpA (toxin co-regulated pilus) production and DNA binding; however, toxT activation by TcpP-H93L was restored in the presence of ToxR, suggesting that ToxR can provide the promoter recognition function for toxT activation. Two additional mutant derivatives, TcpP-W68L and TcpP-R86A, failed to activate toxT or direct toxin and TcpA production in the presence or absence of ToxR. Both TcpP-W68L and TcpP-R86A, like TcpP-H93L, were defective for DNA binding. Finally, a ToxR mutant derivative, ToxR-G80S, served to separate the different roles of ToxR on different promoters. Although ToxR-G80S was inefficient at activating the ompU promoter in V. cholerae (ompU encodes an outer membrane porin regulated by ToxR), it was fully capable of activating the toxT promoter. These data suggest that ToxR is not a direct activator in the toxT expression system but, instead, enhances the activity of TcpP, perhaps by recruiting it to the toxT promoter under conditions in which expression levels of TcpP are too low for it to activate toxT efficiently on its own.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Oct
pubmed:issn
0950-382X
pubmed:author
pubmed:issnType
Print
pubmed:volume
38
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
67-84
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
The Vibrio cholerae ToxR/TcpP/ToxT virulence cascade: distinct roles for two membrane-localized transcriptional activators on a single promoter.
pubmed:affiliation
Unit for Laboratory Animal Medicine, and Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI 48109-0614, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.