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So I know that several different environmental signals, such as pH, bile, and temperature, regulate virulence gene expression in V. cholerae. Specifically, they control expression of the genes encoding the toxin co- regulated pilus (TCP) and cholera toxin (CT).

But what are the two key (main) transcriptional regulatory proteins that control TCP and CT expression, and where is each localized in the bacterial cell? Also, what does each protein do?

I know that the proteins involved are ToxR/S, ToxP/H, and ToxT, but I don't know which are the MAIN ones and why...

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Virulence factors of V cholerae is regulated by a hierarchical regulatory system. The proteins you listed in your question are part of this system. In this paper it is said that the actual activation of TCP and CT is done by ToxT- thus this could be your "key" protein. I'd also like to note that just because these are two different genes does not necessarily mean that there has to be two key factors - this assumption of yours if a bit faulty.

Also because the activation of the virulence factors are regulated by a chain of events, each link is can be considered equally important, because if one of them fails the whole chain is dysfunctional.

The short description of the regulatory chain taken from the article linked above:

The ToxR regulon is a hierarchical regulatory system that activates virulence gene expression in response to environmental stimuli (2). Induction of the ToxR regulon starts with activation of tcpPH expression by AphA and AphB (4, 5). AphA and AphB are cytoplasmic regulatory proteins that cooperatively bind to the tcpPH promoter and activate its expression. Once TcpP is produced, it functions in conjunction with ToxR to activate toxT expression. TcpP and ToxR are structurally similar membrane-bound DNA binding proteins that are thought to modulate gene expression in response to external stimuli (6, 7). When stimulated, TcpP and ToxR bind to the toxT promoter and activate its expression. ToxT then directly activates the expression of the genes that encode for CT and TCP production (8).

This paper might also be helpful. Unfortunately I could only access the full text this way (through researchgate):

Regulation of virulence in Vibrio cholerae: the ToxR regulon. Brandon Childers, Karl E Klose Future Microbiology (Impact Factor: 4.02). 07/2007; 2(3):335-44. DOI: 10.2217/17460913.2.3.335

This paper provides details of regulation and mechanism of ToxT.

ToxT regulation

Taken from the third linked paper. As you can see the final activator of CT (termed CTX in the picture) and TCP is indeed the ToxT, but as I wrote it earlier each link in the chain is important.

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