title: Quantitative Map of β-Lactone-Induced Virulence Regulation.
abstract: β-Lactones have recently been introduced as the first selective ClpP inhibitors that attenuate virulence of both sensitive Staphylococcus aureus and multiresistant strains (MRSA). Although previous knockout studies showed that ClpP is essential for S. aureus alpha-toxin production, a link between β-lactone inhibition and molecular virulence mechanisms has been lacking so far. We here perform a chemical-proteomic approach to elucidate antivirulence pathways. First, we demonstrate by gel-free activity-based protein profiling that ClpP is the predominant target of β-lactones. Only a few off-targets were discovered, which, unlike ClpP, were not involved in the reduction of alpha-toxin expression. Second, in-depth mechanistic insight was provided by a full proteomic comparison between lactone treated and untreated S. aureus cells. Quantitative mass-spectrometric analysis revealed increased repressor of toxin (Rot) levels and a corresponding down-regulation of α-toxin, providing the first direct connection between the lactone-dependent phenotype and a corresponding cellular mechanism. By building up a quantitative virulence regulation network, we visualize the impact of ClpP inhibition in a systems biology context. Interestingly, a lack of in vitro Rot degradation by either ClpXP or ClpCP calls either for a proteolysis mechanism with yet unknown adaptor proteins or for an indirect mode of action that may involve ClpX-mediated RNA signaling and feedback circuits.
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Joanna
authors.initials: J
authors.lastname: Krysiak
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Stahl
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Jan
authors.initials: J
authors.lastname: Vomacka
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Fetzer
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Lakemeyer
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Fux
authors.affiliation: Department of Chemistry, Chair of Organic Chemistry II, Center for Integrated Protein Science (CIPSM), Technische Universität München , Lichtenbergstrasse 4, D-85747 Garching, Germany.
authors.forename: Stephan A
authors.identifier: 0000-0002-9400-906X
authors.initials: SA
authors.lastname: Sieber
mesh_terms: D001426:Bacterial Proteins / Q000032:analysis / Q000235:genetics; D001427:Bacterial Toxins / Q000235:genetics; D049071:Endopeptidase Clp / Q000187:drug effects; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D007783:Lactones / Q000494:pharmacology*; D040901:Proteomics; D012097:Repressor Proteins / Q000032:analysis; D013211:Staphylococcus aureus / Q000472:pathogenicity*; D049490:Systems Biology; D014774:Virulence / Q000187:drug effects*
chemical_list: D001426:Bacterial Proteins; D001427:Bacterial Toxins; D007783:Lactones; D012097:Repressor Proteins; C408643:rot protein, Staphylococcus aureus; D049071:Endopeptidase Clp
keywords: Staphylococcus aureus ClpP; activity-based protein profiling; antivirulence; chemical knockout; proteomics