Regulated intramembrane proteolysis of the virulence activator TcpP in Vibrio cholerae is initiated by the tail-specific protease (Tsp).

Vibrio cholerae uses a multiprotein transcriptional regulatory cascade to control expression of virulence factors cholera toxin and toxin-co-regulated pilus. Two proteins in this cascade are ToxR and TcpP - unusual membrane-localized transcription factors with relatively undefined periplasmic domains and transcription activator cytoplasmic domains. TcpP and ToxR function with each other and two other membrane-localized proteins, TcpH and ToxS, to activate transcription of toxT, encoding the direct activator of toxin and pilus genes. Under some conditions, TcpP is degraded in a two-step proteolytic pathway known as regulated intramembrane proteolysis (RIP), thereby inactivating the cascade. The second step in this proteolytic pathway involves the zinc metalloprotease YaeL; V. cholerae cells lacking YaeL accumulate a truncated yet active form of TcpP termed TcpP*. We hypothesized that a protease acting prior to YaeL degrades TcpP to TcpP*, which is the substrate of YaeL. In this study, we demonstrate that a C-terminal protease called Tsp degrades TcpP to form TcpP*, which is then acted upon by YaeL. We present evidence that TcpH and Tsp serve to protect full-length TcpP from spurious proteolysis by YaeL. Cleavage by Tsp occurs in the periplasmic domain of TcpP and requires residues TcpPA172 and TcpPI174 for wild-type activity.