title: Two-component system VicRK regulates functions associated with establishment of Streptococcus sanguinis in biofilms.
abstract: Streptococcus sanguinis is a commensal pioneer colonizer of teeth and an opportunistic pathogen of infectious endocarditis. The establishment of S. sanguinis in host sites likely requires dynamic fitting of the cell wall in response to local stimuli. In this study, we investigated the two-component system (TCS) VicRK in S. sanguinis (VicRKSs), which regulates genes of cell wall biogenesis, biofilm formation, and virulence in opportunistic pathogens. A vicK knockout mutant obtained from strain SK36 (SKvic) showed slight reductions in aerobic growth and resistance to oxidative stress but an impaired ability to form biofilms, a phenotype restored in the complemented mutant. The biofilm-defective phenotype was associated with reduced amounts of extracellular DNA during aerobic growth, with reduced production of H2O2, a metabolic product associated with DNA release, and with inhibitory capacity of S. sanguinis competitor species. No changes in autolysis or cell surface hydrophobicity were detected in SKvic. Reverse transcription-quantitative PCR (RT-qPCR), electrophoretic mobility shift assays (EMSA), and promoter sequence analyses revealed that VicR directly regulates genes encoding murein hydrolases (SSA_0094, cwdP, and gbpB) and spxB, which encodes pyruvate oxidase for H2O2 production. Genes previously associated with spxB expression (spxR, ccpA, ackA, and tpK) were not transcriptionally affected in SKvic. RT-qPCR analyses of S. sanguinis biofilm cells further showed upregulation of VicRK targets (spxB, gbpB, and SSA_0094) and other genes for biofilm formation (gtfP and comE) compared to expression in planktonic cells. This study provides evidence that VicRKSs regulates functions crucial for S. sanguinis establishment in biofilms and identifies novel VicRK targets potentially involved in hydrolytic activities of the cell wall required for these functions.
authors.affiliation: Department of Oral Diagnosis, Piracicaba Dental School, University of Campinas, Piracicaba, Sao Paulo, Brazil.
authors.forename: Julianna J
authors.initials: JJ
authors.lastname: Moraes
authors.affiliation: Department of Oral Diagnosis, Piracicaba Dental School, University of Campinas, Piracicaba, Sao Paulo, Brazil.
authors.forename: Rafael N
authors.initials: RN
authors.lastname: Stipp
authors.affiliation: Department of Oral Diagnosis, Piracicaba Dental School, University of Campinas, Piracicaba, Sao Paulo, Brazil.
authors.forename: Erika N
authors.initials: EN
authors.lastname: Harth-Chu
authors.affiliation: Department of Oral Diagnosis, Piracicaba Dental School, University of Campinas, Piracicaba, Sao Paulo, Brazil.
authors.forename: Tarsila M
authors.initials: TM
authors.lastname: Camargo
authors.affiliation: Department of Oral Diagnosis, Piracicaba Dental School, University of Campinas, Piracicaba, Sao Paulo, Brazil.
authors.forename: José F
authors.initials: JF
authors.lastname: Höfling
authors.affiliation: Department of Oral Diagnosis, Piracicaba Dental School, University of Campinas, Piracicaba, Sao Paulo, Brazil rmgraner@fop.unicamp.br.
authors.forename: Renata O
authors.initials: RO
authors.lastname: Mattos-Graner
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D018441:Biofilms / Q000254:growth & development*; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D055786:Gene Knockout Techniques; D005816:Genetic Complementation Test; D060888:Real-Time Polymerase Chain Reaction; D013298:Streptococcus sanguis / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: D001426:Bacterial Proteins; C119939:YycF protein, Bacteria