BACKGROUND: The methicillin-resistant Staphylococcus aureus (MRSA) surface protein Pls (plasmin sensitive) reduces adhesion to host proteins and cellular invasiveness by an unknown mechanism that requires Pls expression. Here, we tested the effect of Pls expression using different pls-negative backgrounds. METHODS: Three pls-negative strains (the methicillin-susceptible Staphylococcus aureus strains Cowan I and 6850 and the MRSA strain ST239-635/93, which harbors staphylococcal cassette chromosome [SCC] mec type III) were transformed. Constructs used were full-length pls (pPLS4), pls-DeltaLPDTG (no sortase motif; pPLS5), and pls-DeltaSD (no serine-aspartic acid [SD] repeats; pPLS6). Adherence, invasiveness, gene expression, and surface expression were quantified by photometry, flow cytometry, real-time reverse-transcription polymerase chain reaction, and a modified enzyme-linked immunosorbent assay, respectively. RESULTS: In Pls-expressing strains (those with pPLS4), adherence to immobilized fibronectin (Fn) and binding of soluble Fn was reduced by approximately 20% and approximately 25%, respectively. Invasion of 293 cells and EA.hy 926 cells was reduced by up to 85%. Surprisingly, transcription of fnbA and spa was up-regulated, but transcription of clfA and hla was down-regulated. Pls and Fn-binding protein (FnBP) surface expression was increased. Competition with purified FnBPA, but not with Pls, reduced invasiveness by approximately 90%. The invasiveness of 6850 (pPLS5) and of 6850 (pPLS6) was reduced by only approximately 20% and approximately 15%, respectively. CONCLUSION: Expression of cell wall-anchored Pls reduces adherence and invasiveness independently of the MRSA/SCCmec background. This occurs despite early up-regulation of fnbA transcription and FnBP surface expression. Thus, Pls acts by steric hindrance rather than another mechanism.
BACKGROUND: The methicillin-resistant Staphylococcus aureus (MRSA) surface protein Pls (plasmin sensitive) reduces adhesion to host proteins and cellular invasiveness by an unknown mechanism that requires Pls expression. Here, we tested the effect of Pls expression using different pls-negative backgrounds. METHODS: Three pls-negative strains (the methicillin-susceptible Staphylococcus aureus strains Cowan I and 6850 and the MRSA strain ST239-635/93, which harbors staphylococcal cassette chromosome [SCC] mec type III) were transformed. Constructs used were full-length pls (pPLS4), pls-DeltaLPDTG (no sortase motif; pPLS5), and pls-DeltaSD (no serine-aspartic acid [SD] repeats; pPLS6). Adherence, invasiveness, gene expression, and surface expression were quantified by photometry, flow cytometry, real-time reverse-transcription polymerase chain reaction, and a modified enzyme-linked immunosorbent assay, respectively. RESULTS: In Pls-expressing strains (those with pPLS4), adherence to immobilized fibronectin (Fn) and binding of soluble Fn was reduced by approximately 20% and approximately 25%, respectively. Invasion of 293 cells and EA.hy 926 cells was reduced by up to 85%. Surprisingly, transcription of fnbA and spa was up-regulated, but transcription of clfA and hla was down-regulated. Pls and Fn-binding protein (FnBP) surface expression was increased. Competition with purified FnBPA, but not with Pls, reduced invasiveness by approximately 90%. The invasiveness of 6850 (pPLS5) and of 6850 (pPLS6) was reduced by only approximately 20% and approximately 15%, respectively. CONCLUSION: Expression of cell wall-anchored Pls reduces adherence and invasiveness independently of the MRSA/SCCmec background. This occurs despite early up-regulation of fnbA transcription and FnBP surface expression. Thus, Pls acts by steric hindrance rather than another mechanism.
Authors: P Balamurugan; V Praveen Krishna; D Bharath; Raajaraam Lavanya; Pothiappan Vairaprakash; S Adline Princy Journal: Front Microbiol Date: 2017-07-11 Impact factor: 5.640
Authors: Heidi A Crosby; Patrick M Schlievert; Joseph A Merriman; Jessica M King; Wilmara Salgado-Pabón; Alexander R Horswill Journal: PLoS Pathog Date: 2016-05-04 Impact factor: 6.823