CvfA protein and polynucleotide phosphorylase act in an opposing manner to regulate Staphylococcus aureus virulence.
We previously identified CvfA (SA1129) as a Staphylococcus aureus virulence factor using a silkworm infection model. S. aureus cvfA-deleted mutants exhibit decreased expression of the agr locus encoding a positive regulator of hemolysin genes and decreased hemolysin production. CvfA protein hydrolyzes a 2',3'-cyclic phosphodiester bond at the RNA 3' terminus, producing RNA with a 3'-phosphate (3'-phosphorylated RNA, RNA with a 3'-phosphate). Here, we report that the cvfA-deleted mutant phenotype (decreased agr expression and hemolysin production) was suppressed by disrupting pnpA-encoding polynucleotide phosphorylase (PNPase) with 3'- to 5'-exonuclease activity. The suppression was blocked by introducing a pnpA-encoding PNPase with exonuclease activity but not by a pnpA-encoding mutant PNPase without exonuclease activity. Therefore, loss of PNPase exonuclease activity suppressed the cvfA-deleted mutant phenotype. Purified PNPase efficiently degraded RNA with 2',3'-cyclic phosphate at the 3' terminus (2',3'-cyclic RNA), but it inefficiently degraded 3'-phosphorylated RNA. These findings indicate that 3'-phosphorylated RNA production from 2',3'-cyclic RNA by CvfA prevents RNA degradation by PNPase and contributes to the expression of agr and hemolysin genes. We speculate that in the cvfA-deleted mutant, 2',3'-cyclic RNA is not converted to the 3'-phosphorylated form and is efficiently degraded by PNPase, resulting in the loss of RNA essential for expressing agr and hemolysin genes, whereas in the cvfA/pnpA double-disrupted mutant, 2',3'-cyclic RNA is not degraded by PNPase, leading to hemolysin production. These findings suggest that CvfA and PNPase competitively regulate RNA degradation essential for S. aureus virulence.
From the Laboratory of Microbiology, Graduate School of Pharmaceutical Sciences, The University of Tokyo, 3-1, 7-chome, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
Shunsuke
S
Numata
Makiko
M
Nagata
Han
H
Mao
Kazuhisa
K
Sekimizu
Chikara
C
Kaito
D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D012831:Bombyx / Q000382:microbiology; D005260:Female; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism; D054884:Host-Pathogen Interactions; D006801:Humans; D051379:Mice; D010727:Phosphoric Diester Hydrolases / Q000235:genetics / Q000378:metabolism*; D011117:Polyribonucleotide Nucleotidyltransferase / Q000235:genetics / Q000378:metabolism*; D012329:RNA, Bacterial / Q000235:genetics / Q000378:metabolism; D013203:Staphylococcal Infections / Q000378:metabolism / Q000382:microbiology*; D013211:Staphylococcus aureus / Q000472:pathogenicity* / Q000502:physiology; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D006460:Hemolysin Proteins; D012329:RNA, Bacterial; D037521:Virulence Factors; D011117:Polyribonucleotide Nucleotidyltransferase; D010727:Phosphoric Diester Hydrolases
Bacterial Genetics; Bacterial Pathogenesis; RNA Metabolism; RNA Modification; RNA Processing; Ribonuclease; Staphylococcus aureus