Global Regulatory Impact of ClpP Protease ofStaphylococcus aureuson Regulons Involved in Virulence, Oxidative Stress Response, Autolysis, and DNA Repair

Abstract

Staphylococcus aureus is an important pathogen, causing a wide range of infections including sepsis, wound infections, pneumonia, and catheter-related infections. In several pathogens ClpP proteases were identified by in vivo expression technologies to be important for virulence. Clp proteolytic complexes are responsible for adaptation to multiple stresses by degrading accumulated and misfolded proteins. In this report clpP, encoding the proteolytic subunit of the ATP-dependent Clp protease, was deleted, and gene expression of ⌬clpP was determined by global transcriptional analysis using DNA-microarray technology. The transcriptional profile reveals a strong regulatory impact of ClpP on the expression of genes encoding proteins that are involved in the pathogenicity of S. aureus and adaptation of the pathogen to several stresses. Expression of the agr system and agr-dependent extracellular virulence factors was diminished. Moreover, the loss of clpP leads to a complete transcriptional derepression of genes of the CtsR-and HrcA-controlled heat shock regulon and a partial derepression of genes involved in oxidative stress response, metal homeostasis, and SOS DNA repair controlled by PerR, Fur, MntR, and LexA. The levels of transcription of genes encoding proteins involved in adaptation to anaerobic conditions potentially regulated by an Fnr-like regulator were decreased. Furthermore, the expression of genes whose products are involved in autolysis was deregulated, leading to enhanced autolysis in the mutant. Our results indicate a strong impact of ClpP proteolytic activity on virulence, stress response, and physiology in S. aureus.The Clp proteases were first identified in Escherichia coli and consist of an ATPase specificity factor (ClpA or ClpX in E. coli; ClpX, ClpC, or ClpE in Bacillus subtilis) and a proteolytic domain (ClpP) that contains a consensus serine protease active site (33). In E. coli, ClpP-mediated proteolysis is regulated by heat shock and removes abnormal proteins that accumulate during stress conditions, recycles amino acids from nonessential proteins during starvation, and contributes to the clearance of truncated peptides from stalled ribosomes by the SsrAtagging system (34, 65). Moreover, it has been shown that Clp proteases play a significant role in certain processes regulating cellular functions via proteolysis (33,36,45). For example, in E. coli ClpXP is involved in degradation of regulator proteins including the alternative sigma factor SigS, the UmuD SOS protein, and different phage proteins (23,31,34 (44,69,73,74). In B. subtilis Clp-specific target proteins were recognized which are involved in peptidoglycan synthesis, competence, sporulation, and heat shock regulation (30,48,50).In addition, several studies indicate that ClpP proteolytic activity is critical for virulence of pathogenic bacteria, including S. enterica serovar Typhimurium, Streptococcus pneumoniae, Listeria monocytogenes, and Staphylococcus aureus (26,29,37,58,63,73,75). Interestingly, ClpP seems to be essential for s...

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