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- W3031719606 abstract "ObjectiveTo construct a ciaR gene-knockout (ΔciaR) mutant of Streptococcus pneumoniae (S.pneumoniae) and to investigate the effects of CiaR in CiaH/CiaR, a streptococcal two-component signal-transducing system, on the expression of genes encoding penicillin-binding proteins (pbps genes) and cia-dependent small RNAs (csRNAs).MethodsElectrophoretic mobility shift assay (ESMA) was performed to detect the recombinant CiaR (rCiaR)-binding pbps genes. A suicide plasmid pEVP3ciaR for ciaR gene knockout was constructed and then a ΔciaR mutant was obtained through homologous recombination and insertion inactivation of the suicide plasmid, and screening with chloromycin. The mutant was identified using PCR and sequencing analysis. E-test was used to detect the minimal inhibitory concentrations (MIC) of penicillin (PCN) and cefotaxime (CTX) against S. pneumoniae strains. Changes and differences in the expression of pbps genes and csRNAs in the ΔciaR mutant and its wild-type strain before and after treatment with 1/4 MIC of PCN or CTX were detected using real-time quantitative RT-PCR.ResultsThe rCiaR could bind to the promoter regions in pbp1a, pbp1b and pbp2b genes of S. pneumoniae. The ciaR gene in ΔciaR mutant was inactivated by insertion according to the results of PCR and sequencing analysis. After treatment with 1/4 MIC of PCN or CTX, the expression of pbps genes at mRNA level (pbps-mRNAs) in the ΔciaR mutant was significantly increased (P<0.05), but the levels of csRNAs were significantly decreased (P<0.05); whereas a significantly decreased pbps-mRNAs (P<0.05) and increased csRNAs (P<0.05) were observed in its wild-type strain. The result of E-test showed that the MICs of PCN and CTX against ΔciaR mutant were increased by 250-fold as compared with those against its wild-type strain.ConclusionThe CiaR can enhance the drug resistance of S. pneumoniae to PCN and CTX through down-regulating the expression of PBP1a, PBP1b and PBP2b and up-regulating the expression of csRNAs to inhibit the expression of PBPs.Key words: Streptococcus pneumoniae; Drug resistance; Response regulation protein/CiaR; Gene knockout; PBPs/csRNAs" @default.
- W3031719606 created "2020-06-05" @default.
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- W3031719606 date "2017-01-31" @default.
- W3031719606 modified "2023-09-24" @default.
- W3031719606 title "Roles of Streptococcus pneumoniae CiaR in regulating the expression of pbps genes and csRNAs and its correlation with drug resistance" @default.
- W3031719606 doi "https://doi.org/10.3760/cma.j.issn.0254-5101.2017.01.008" @default.
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