title: Proteomic and transcriptomic profiling of Staphylococcus aureus surface LPXTG-proteins: correlation with agr genotypes and adherence phenotypes.
abstract: Staphylococcus aureus infections involve numerous adhesins and toxins, which expression depends on complex regulatory networks. Adhesins include a family of surface proteins covalently attached to the peptidoglycan via a conserved LPXTG motif. Here we determined the protein and mRNA expression of LPXTG-proteins of S. aureus Newman in time-course experiments, and their relation to fibrinogen adherence in vitro. Experiments were performed with mutants in the global accessory-gene regulator (agr), surface protein A (Spa), and fibrinogen-binding protein A (ClfA), as well as during growth in iron-rich or iron-poor media. Surface proteins were recovered by trypsin-shaving of live bacteria. Released peptides were analyzed by liquid chromatography coupled to tandem mass-spectrometry. To unambiguously identify peptides unique to LPXTG-proteins, the analytical conditions were refined using a reference library of S. aureus LPXTG-proteins heterogeneously expressed in surrogate Lactococcus lactis. Transcriptomes were determined by microarrays. Sixteen of the 18 LPXTG-proteins present in S. aureus Newman were detected by proteomics. Nine LPXTG-proteins showed a bell-shape agr-like expression that was abrogated in agr-negative mutants including Spa, fibronectin-binding protein A (FnBPA), ClfA, iron-binding IsdA, and IsdB, immunomodulator SasH, functionally uncharacterized SasD, biofilm-related SasG and methicillin resistance-related FmtB. However, only Spa and SasH modified their proteomic and mRNA profiles in parallel in the parent and its agr- mutant, whereas all other LPXTG-proteins modified their proteomic profiles independently of their mRNA. Moreover, ClfA became highly transcribed and active in fibrinogen-adherence tests during late growth (24 h), whereas it remained poorly detected by proteomics. On the other hand, iron-regulated IsdA-B-C increased their protein expression by >10-times in iron-poor conditions. Thus, proteomic, transcriptomic, and adherence-phenotype demonstrated differential profiles in S. aureus. Moreover, trypsin peptide signatures suggested differential protein domain exposures in various environments, which might be relevant for anti-adhesin vaccines. A comprehensive understanding of the S. aureus physiology should integrate all three approaches.
authors: Department of Fundamental Microbiology, University of Lausanne, Lausanne, Switzerland.
authors: Mathilde
authors: M
authors: Ythier
authors: Grégory
authors: G
authors: Resch
authors: Patrice
authors: P
authors: Waridel
authors: Alexandre
authors: A
authors: Panchaud
authors: Aurélie
authors: A
authors: Gfeller
authors: Paul
authors: P
authors: Majcherczyk
authors: Manfredo
authors: M
authors: Quadroni
authors: Philippe
authors: P
authors: Moreillon
mesh_terms: D001422:Bacterial Adhesion / Q000187:drug effects / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D030562:Databases, Protein; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005838:Genotype; D007501:Iron / Q000494:pharmacology; D007700:Kinetics; D017045:Lactococcus / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D050296:Microbial Viability / Q000187:drug effects; D009154:Mutation / Q000235:genetics*; D019151:Peptide Library; D010455:Peptides / Q000378:metabolism; D010641:Phenotype; D040901:Proteomics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D013211:Staphylococcus aureus / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; D019151:Peptide Library; D010455:Peptides; D012333:RNA, Messenger; D007501:Iron; D014357:Trypsin