Distribution and expression of the ZmpA metalloprotease in the Burkholderia cepacia complex.
The distribution of the metalloprotease gene zmpA was determined among strains of the Burkholderia cepacia complex (Bcc). The zmpA gene was present in B. cepacia, B. cenocepacia, B. stabilis, B. ambifaria and B. pyrrocinia but absent from B. multivorans, B. vietnamiensis, B. dolosa, and B. anthina. The presence of zmpA generally correlated with extracellular proteolytic activity with the exception of five strains, which had zmpA but had no detectable proteolytic activity when skim milk agar was used as a substrate (zmpA protease deficient). Western immunoblot experiments with anti-ZmpA antibodies suggest that the zmpA protease-deficient strains do not secrete or accumulate detectable ZmpA. Transcriptional zmpA::lacZ fusions were introduced in selected strains of the Bcc. zmpA::lacZ was expressed in all strains, but expression was generally lower in the zmpA protease-deficient strains than in the zmpA protease-proficient strains. Quantitative reverse transcriptase real-time PCR demonstrated that zmpA protease-deficient strains did express zmpA mRNA, although at various levels. ZmpA has previously been shown to be positively regulated by the CepIR quorum-sensing system. Addition of exogenous AHLs did not restore extracellular protease production to any of the zmpA protease-deficient strains; however, introduction of cepR in trans complemented protease activity in two of five strains. Extracellular proteolytic activity was restored by the presence of zmpA in trans in two of the five strains. These studies suggest that although some strains of the Bcc contain the zmpA gene, multiple factors may influence its expression.
Gingues
S
S
Department of Microbiology and Infectious Diseases, Faculty of Medicine, University of Calgary Health Sciences Centre, Calgary, Alberta T2N 4N1, Canada.
Kooi
C
C
Visser
M B
MB
Subsin
B
B
Sokol
P A
PA
D015107:4-Butyrolactone / Q000031:analogs & derivatives / Q000502:physiology; D000818:Animals; D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D015153:Blotting, Western; D042602:Burkholderia cepacia complex / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D002364:Caseins / Q000378:metabolism; D015870:Gene Expression; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D008025:Ligases / Q000235:genetics / Q000378:metabolism; D008666:Metalloendopeptidases / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D012329:RNA, Bacterial / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D002364:Caseins; D012329:RNA, Bacterial; D012333:RNA, Messenger; C088386:homoserine lactone; D008666:Metalloendopeptidases; C502465:ZmpA protein, Burkholderia cenocepacia; D008025:Ligases; C449816:N-acylhomoserine lactone synthase; D015107:4-Butyrolactone