Analysis of the virulence-associated RevSR two-component signal transduction system of Clostridium perfringens.
Clostridium perfringens is a Gram-positive, anaerobic, spore-forming bacterium that causes human gas gangrene (clostridial myonecrosis) and food poisoning. Early studies showed that virulence was regulated by the VirSR two-component signal transduction system. However, our identification of the RevR orphan response regulator indicated that more than one system was involved in controlling virulence. To further characterize this virulence-associated regulator, gel mobility shift experiments, coupled with DNase I footprinting, were used to identify the RevR DNA binding sequence. Bioinformatics analysis suggested that an orphan sensor histidine kinase, CPE1757 (renamed RevS), was the cognate sensor of RevR. Interaction between RevS and RevR was demonstrated by use of a bacterial two-hybrid system and validated by protein-protein interaction studies using biolayer interferometry. To assess the involvement of RevS in virulence regulation, the revS gene was inactivated by Targetron insertion. When isogenic wild-type, revS and complemented revS strains were tested in a mouse myonecrosis model, the revS mutant was found to be attenuated in virulence, which was similar to the attenuation observed previously with the revR mutant. However, transcriptional analysis of selected RevR-regulated genes in the revS mutant revealed a different pattern of expression to a revR mutant, suggesting that the RevSR system is more complex than originally thought. Taken together, the results have led to the identification and characterization of the two essential parts of a new regulatory network that is involved in the regulation of virulence in C. perfringens.
Infection and Immunity Program, Biomedicine Discovery Institute and Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia.
Jackie K
JK
Cheung
Infection and Immunity Program, Biomedicine Discovery Institute and Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia.
Jessica A
JA
Wisniewski
Infection and Immunity Program, Biomedicine Discovery Institute and Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia.
Vicki M
VM
Adams
Protein Production Unit, Monash University, Clayton, Victoria 3800, Australia.
Noelene S
NS
Quinsey
Infection and Immunity Program, Biomedicine Discovery Institute and Department of Microbiology, Monash University, Clayton, Victoria 3800, Australia. Electronic address: julian.rood@monash.edu.
Julian I
JI
Rood
D000818:Animals; D001665:Binding Sites; D003016:Clostridium perfringens / Q000235:genetics / Q000502:physiology*; D018983:DNA Footprinting; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D005260:Female; D015964:Gene Expression Regulation, Bacterial; D055786:Gene Knockout Techniques; D005816:Genetic Complementation Test; D000071677:Histidine Kinase / Q000235:genetics / Q000378:metabolism*; D008807:Mice, Inbred BALB C; D016254:Mutagenesis, Insertional; D011485:Protein Binding; D025941:Protein Interaction Mapping; D015398:Signal Transduction; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014774:Virulence
D004269:DNA, Bacterial; D014157:Transcription Factors; D000071677:Histidine Kinase
Clostridium perfringens; Gene regulation; Two-component system; Virulence