Dataset Information

Evidence of a bacterial receptor for lysozyme: binding of lysozyme to the anti-? factor RsiV controls activation of the ecf ? factor ?V.

ABSTRACT: ? factors endow RNA polymerase with promoter specificity in bacteria. Extra-Cytoplasmic Function (ECF) ? factors represent the largest and most diverse family of ? factors. Most ECF ? factors must be activated in response to an external signal. One mechanism of activation is the stepwise proteolytic destruction of an anti-? factor via Regulated Intramembrane Proteolysis (RIP). In most cases, the site-1 protease required to initiate the RIP process directly senses the signal. Here we report a new mechanism in which the anti-? factor rather than the site-1 protease is the sensor. We provide evidence suggesting that the anti-? factor RsiV is the bacterial receptor for the innate immune defense enzyme, lysozyme. The site-1 cleavage site is similar to the recognition site of signal peptidase and cleavage at this site is required for ?V activation in Bacillus subtilis. We reconstitute site-1 cleavage in vitro and demonstrate that it requires both signal peptidase and lysozyme. We demonstrate that the anti-? factor RsiV directly binds to lysozyme and muramidase activity is not required for ?V activation. We propose a model in which the binding of lysozyme to RsiV activates RsiV for signal peptidase cleavage at site-1, initiating proteolytic destruction of RsiV and activation of ?V. This suggests a novel mechanism in which conformational change in a substrate controls the cleavage susceptibility for signal peptidase. Thus, unlike other ECF ? factors which require regulated intramembrane proteolysis for activation, the sensor for ?V activation is not the site-1 protease but the anti-? factor.

SUBMITTER: Hastie JL

PROVIDER: S-EPMC4183432 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Evidence of a bacterial receptor for lysozyme: binding of lysozyme to the anti-σ factor RsiV controls activation of the ecf σ factor σV.

Hastie Jessica L JL Williams Kyle B KB Sepúlveda Carolina C Houtman Jon C JC Forest Katrina T KT Ellermeier Craig D CD

PLoS genetics 20141002 10

σ factors endow RNA polymerase with promoter specificity in bacteria. Extra-Cytoplasmic Function (ECF) σ factors represent the largest and most diverse family of σ factors. Most ECF σ factors must be activated in response to an external signal. One mechanism of activation is the stepwise proteolytic destruction of an anti-σ factor via Regulated Intramembrane Proteolysis (RIP). In most cases, the site-1 protease required to initiate the RIP process directly senses the signal. Here we report a new ...[more]

PMID: 25275625

Similar Datasets

Project description:Clostridioides (Clostridium) difficile is one of the leading causes of nosocomial diarrhea. Lysozyme is a common host defense against many pathogenic bacteria. C. difficile exhibits high levels of lysozyme resistance, which is due in part to the extracytoplasmic functioning (ECF) σ factor, σV. It has been previously demonstrated that genes regulated by σV are responsible for peptidoglycan modifications that provide C. difficile with high lysozyme resistance. σV is not unique to C. difficile however, and its role in lysozyme resistance and its mechanism of activation has been well characterized in Bacillus subtilis where the anti-σ, RsiV, sequesters σV until lysozyme directly binds to RsiV, activating σV. However, it remains unclear if the mechanism of σV activation is similar in C. difficile. Here, we investigated how activation of σV is controlled in C. difficile by lysozyme. We found that C. difficile RsiV was degraded in the presence of lysozyme. We also found that disruption of a predicted signal peptidase cleavage site blocked RsiV degradation and σV activation, indicating that the site-1 protease is likely a signal peptidase. We also identified a conserved site-2 protease, RasP, that was required for site-2 cleavage of RsiV and σV activation in response to lysozyme. Combined with previous work showing RsiV directly binds lysozyme, these data suggested that RsiV directly binds lysozyme in C. difficile, which leads to RsiV destruction via cleavage at site-1 by signal peptidase and then at site-2 by RasP, ultimately resulting in σV activation and increased resistance to lysozyme. IMPORTANCE Clostridioides difficile is a major cause of hospital-acquired diarrhea and represents an urgent concern due to the prevalence of antibiotic resistance and the rate of recurrent infections. We previously showed that σV and the regulon under its control were involved in lysozyme resistance. We have also shown in B. subtilis that the anti-σ RsiV acts as a direct sensor for lysozyme. which results in the destruction of RsiV and activation of σV. Here, we described the proteases required for degradation of RsiV in C. difficile in response to lysozyme. Our data indicated that the mechanism is highly conserved between B. subtilis and C. difficile.

Project description:Extracytoplasmic function (ECF) ? factors are a diverse family of alternative ? factors that allow bacteria to sense and respond to changes in the environment. ?V is an ECF ? factor found primarily in low-GC Gram-positive bacteria and is required for lysozyme resistance in several opportunistic pathogens. In the absence of lysozyme, ?V is inhibited by the anti-? factor RsiV. In response to lysozyme, RsiV is degraded via the process of regulated intramembrane proteolysis (RIP). RIP is initiated by cleavage of RsiV at site 1, which allows the intramembrane protease RasP to cleave RsiV within the transmembrane domain at site 2 and leads to activation of ?V Previous work suggested that RsiV is cleaved by signal peptidase at site 1. Here we demonstrate in vitro that signal peptidase is sufficient for cleavage of RsiV only in the presence of lysozyme and provide evidence that multiple Bacillus subtilis signal peptidases can cleave RsiV in vitro This cleavage is dependent upon the concentration of lysozyme, consistent with previous work that showed that binding to RsiV was required for ?V activation. We also show that signal peptidase activity is required for site 1 cleavage of RsiV in vivo Thus, we demonstrate that signal peptidase is the site 1 protease for RsiV.IMPORTANCE Extracytoplasmic function (ECF) ? factors are a diverse family of alternative ? factors that respond to extracellular signals. The ECF ? factor ?V is present in many low-GC Gram-positive bacteria and induces resistance to lysozyme, a component of the innate immune system. The anti-? factor RsiV inhibits ?V activity in the absence of lysozyme. Lysozyme binds RsiV, which initiates a proteolytic cascade leading to destruction of RsiV and activation of ?V This proteolytic cascade is initiated by signal peptidase, a component of the general secretory system. We show that signal peptidase is necessary and sufficient for cleavage of RsiV at site 1 in the presence of lysozyme. This report describes a role for signal peptidase in controlling gene expression.

Dataset Information

Evidence of a bacterial receptor for lysozyme: binding of lysozyme to the anti-? factor RsiV controls activation of the ecf ? factor ?V.

Publications

Evidence of a bacterial receptor for lysozyme: binding of lysozyme to the anti-σ factor RsiV controls activation of the ecf σ factor σV.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets