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Proteolysis of virulence regulator ToxR is associated with entry of Vibrio cholerae into a dormant state.

Almagro-Moreno S, Kim TK, Skorupski K, Taylor RK - PLoS Genet. (2015)

Bottom Line: Strains that can proteolyze ToxR, or do not encode it, lose culturability, experience a change in morphology associated with cells in VBNC, yet remain viable under nutrient limitation at alkaline pH.On the other hand, mutant strains that cannot proteolyze ToxR remain culturable and maintain the morphology of cells in an active state of growth.Overall, our findings provide a link between the proteolysis of a virulence regulator and the entry of a pathogen into an environmentally persistent state.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire, United States of America.

ABSTRACT
Vibrio cholerae O1 is a natural inhabitant of aquatic environments and causes the diarrheal disease, cholera. Two of its primary virulence regulators, TcpP and ToxR, are localized in the inner membrane. TcpP is encoded on the Vibrio Pathogenicity Island (VPI), a horizontally acquired mobile genetic element, and functions primarily in virulence gene regulation. TcpP has been shown to undergo regulated intramembrane proteolysis (RIP) in response to environmental conditions that are unfavorable for virulence gene expression. ToxR is encoded in the ancestral genome and is present in non-pathogenic strains of V. cholerae, indicating it has roles outside of the human host. In this study, we show that ToxR undergoes RIP in V. cholerae in response to nutrient limitation at alkaline pH, a condition that occurs during the stationary phase of growth. This process involves the site-2 protease RseP (YaeL), and is dependent upon the RpoE-mediated periplasmic stress response, as deletion mutants for the genes encoding these two proteins cannot proteolyze ToxR under nutrient limitation at alkaline pH. We determined that the loss of ToxR, genetically or by proteolysis, is associated with entry of V. cholerae into a dormant state in which the bacterium is normally found in the aquatic environment called viable but nonculturable (VBNC). Strains that can proteolyze ToxR, or do not encode it, lose culturability, experience a change in morphology associated with cells in VBNC, yet remain viable under nutrient limitation at alkaline pH. On the other hand, mutant strains that cannot proteolyze ToxR remain culturable and maintain the morphology of cells in an active state of growth. Overall, our findings provide a link between the proteolysis of a virulence regulator and the entry of a pathogen into an environmentally persistent state.

No MeSH data available.


Related in: MedlinePlus

Sequential model for the RIP of ToxR during late stationary phase.(A) In the early stages of colonization, when nutrients are abundant, ToxRS upregulate the expression of genes such as ompU important for growth under these conditions and downregulate the expression of genes such as ompT with roles in environmental survival. (B) During the late stages of colonization, as nutrients become depleted and the environment becomes alkalinized, ToxR is proteolyzed. This occurs due to activation of the σE pathway via sequential degradation of RseA by either DegS or another site-1 protease (1) and RseP (2), which releases RpoE to activate its regulated genes (3). One of these genes may encode a site-1 protease (P) that enters the periplasm (4) and cleaves a periplasmic portion of ToxR, however, at least a second protease/system (P2) appears to be necessary for the site-1 proteolytic event to occur (5). This event is followed by proteolysis of an inner-membrane site of ToxR by RseP (6), which then induces and prevents, respectively, expression of ToxR repressed and activated genes.
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pgen.1005145.g008: Sequential model for the RIP of ToxR during late stationary phase.(A) In the early stages of colonization, when nutrients are abundant, ToxRS upregulate the expression of genes such as ompU important for growth under these conditions and downregulate the expression of genes such as ompT with roles in environmental survival. (B) During the late stages of colonization, as nutrients become depleted and the environment becomes alkalinized, ToxR is proteolyzed. This occurs due to activation of the σE pathway via sequential degradation of RseA by either DegS or another site-1 protease (1) and RseP (2), which releases RpoE to activate its regulated genes (3). One of these genes may encode a site-1 protease (P) that enters the periplasm (4) and cleaves a periplasmic portion of ToxR, however, at least a second protease/system (P2) appears to be necessary for the site-1 proteolytic event to occur (5). This event is followed by proteolysis of an inner-membrane site of ToxR by RseP (6), which then induces and prevents, respectively, expression of ToxR repressed and activated genes.

Mentions: A sequential model for the RIP of ToxR during late stationary phase is shown in Fig 8. In the early stages of colonization, when nutrients are abundant, ToxR upregulates the expression of genes such as ompU and those involved in virulence and downregulates the expression of genes such as ompT with roles in environmental survival. During the late stages of colonization, as nutrients become depleted and the environment becomes alkalinized, ToxR is proteolyzed. This occurs due to activation of the σE pathway via sequential degradation of RseA by either DegS or another site-1 protease and RseP, the site-2 protease, which releases RpoE to activate its regulated genes. It is possible that one of these genes may encode a site-1 protease that cleaves a periplasmic portion of ToxR, however, at least a second protease/system appears to be necessary for the site-1 proteolytic event to occur. Next, RseP cleaves at an intramembrane site within ToxR leading to full proteolysis of the regulator. This process induces and prevents, respectively, expression of ToxR repressed and activated genes, providing an advantage in the environment. The loss of ToxR ultimately leads to entry into dormancy.


Proteolysis of virulence regulator ToxR is associated with entry of Vibrio cholerae into a dormant state.

Almagro-Moreno S, Kim TK, Skorupski K, Taylor RK - PLoS Genet. (2015)

Sequential model for the RIP of ToxR during late stationary phase.(A) In the early stages of colonization, when nutrients are abundant, ToxRS upregulate the expression of genes such as ompU important for growth under these conditions and downregulate the expression of genes such as ompT with roles in environmental survival. (B) During the late stages of colonization, as nutrients become depleted and the environment becomes alkalinized, ToxR is proteolyzed. This occurs due to activation of the σE pathway via sequential degradation of RseA by either DegS or another site-1 protease (1) and RseP (2), which releases RpoE to activate its regulated genes (3). One of these genes may encode a site-1 protease (P) that enters the periplasm (4) and cleaves a periplasmic portion of ToxR, however, at least a second protease/system (P2) appears to be necessary for the site-1 proteolytic event to occur (5). This event is followed by proteolysis of an inner-membrane site of ToxR by RseP (6), which then induces and prevents, respectively, expression of ToxR repressed and activated genes.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4388833&req=5

pgen.1005145.g008: Sequential model for the RIP of ToxR during late stationary phase.(A) In the early stages of colonization, when nutrients are abundant, ToxRS upregulate the expression of genes such as ompU important for growth under these conditions and downregulate the expression of genes such as ompT with roles in environmental survival. (B) During the late stages of colonization, as nutrients become depleted and the environment becomes alkalinized, ToxR is proteolyzed. This occurs due to activation of the σE pathway via sequential degradation of RseA by either DegS or another site-1 protease (1) and RseP (2), which releases RpoE to activate its regulated genes (3). One of these genes may encode a site-1 protease (P) that enters the periplasm (4) and cleaves a periplasmic portion of ToxR, however, at least a second protease/system (P2) appears to be necessary for the site-1 proteolytic event to occur (5). This event is followed by proteolysis of an inner-membrane site of ToxR by RseP (6), which then induces and prevents, respectively, expression of ToxR repressed and activated genes.
Mentions: A sequential model for the RIP of ToxR during late stationary phase is shown in Fig 8. In the early stages of colonization, when nutrients are abundant, ToxR upregulates the expression of genes such as ompU and those involved in virulence and downregulates the expression of genes such as ompT with roles in environmental survival. During the late stages of colonization, as nutrients become depleted and the environment becomes alkalinized, ToxR is proteolyzed. This occurs due to activation of the σE pathway via sequential degradation of RseA by either DegS or another site-1 protease and RseP, the site-2 protease, which releases RpoE to activate its regulated genes. It is possible that one of these genes may encode a site-1 protease that cleaves a periplasmic portion of ToxR, however, at least a second protease/system appears to be necessary for the site-1 proteolytic event to occur. Next, RseP cleaves at an intramembrane site within ToxR leading to full proteolysis of the regulator. This process induces and prevents, respectively, expression of ToxR repressed and activated genes, providing an advantage in the environment. The loss of ToxR ultimately leads to entry into dormancy.

Bottom Line: Strains that can proteolyze ToxR, or do not encode it, lose culturability, experience a change in morphology associated with cells in VBNC, yet remain viable under nutrient limitation at alkaline pH.On the other hand, mutant strains that cannot proteolyze ToxR remain culturable and maintain the morphology of cells in an active state of growth.Overall, our findings provide a link between the proteolysis of a virulence regulator and the entry of a pathogen into an environmentally persistent state.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire, United States of America.

ABSTRACT
Vibrio cholerae O1 is a natural inhabitant of aquatic environments and causes the diarrheal disease, cholera. Two of its primary virulence regulators, TcpP and ToxR, are localized in the inner membrane. TcpP is encoded on the Vibrio Pathogenicity Island (VPI), a horizontally acquired mobile genetic element, and functions primarily in virulence gene regulation. TcpP has been shown to undergo regulated intramembrane proteolysis (RIP) in response to environmental conditions that are unfavorable for virulence gene expression. ToxR is encoded in the ancestral genome and is present in non-pathogenic strains of V. cholerae, indicating it has roles outside of the human host. In this study, we show that ToxR undergoes RIP in V. cholerae in response to nutrient limitation at alkaline pH, a condition that occurs during the stationary phase of growth. This process involves the site-2 protease RseP (YaeL), and is dependent upon the RpoE-mediated periplasmic stress response, as deletion mutants for the genes encoding these two proteins cannot proteolyze ToxR under nutrient limitation at alkaline pH. We determined that the loss of ToxR, genetically or by proteolysis, is associated with entry of V. cholerae into a dormant state in which the bacterium is normally found in the aquatic environment called viable but nonculturable (VBNC). Strains that can proteolyze ToxR, or do not encode it, lose culturability, experience a change in morphology associated with cells in VBNC, yet remain viable under nutrient limitation at alkaline pH. On the other hand, mutant strains that cannot proteolyze ToxR remain culturable and maintain the morphology of cells in an active state of growth. Overall, our findings provide a link between the proteolysis of a virulence regulator and the entry of a pathogen into an environmentally persistent state.

No MeSH data available.


Related in: MedlinePlus