FIG 2.
Phosphorylation levels of CovR and the expression of CovR, rgg, and speB in the wild-type NZ131 strain (emm49 type) and its CovRD53A and CovRT65A mutants. Bacterial strains were cultured to the exponential (for detecting CovR) and stationary (for detecting rgg and speB) phases of growth. RNAs, culture supernatants, and total proteins were collected for RT-qPCR, Western blotting, and Phos-tag Western blot analyses. (A) Levels of phosphorylation and expression of CovR protein in NZ131, CovRD53A mutant, and CovRT65A mutant. CovR∼P, phosphorylated CovR; CovR, nonphosphorylated CovR. Total protein serves as the loading control. (B) Transcription of rgg in NZ131, CovRD53A mutant, and CovRT65A mutant. (C) Transcription of speB and secretion of SpeB protein in NZ131, CovRD53A mutant, and CovRT65A mutant. Biological replicate experiments were performed using three independent preparations. The expression of rgg and speB was normalized to that of gyrA. Wt, wild-type strain; *, P < 0.05.