Fig. 3

Negative regulation of ncgl0887 expression by AtsR. A, B qRT-PCR assay was performed to analyze the expression of ncgl0887 in WT(pXMJ19), ΔatsR(pXMJ19), ΔatsR(pXMJ19-atsR), and ΔatsR(pXMJ19-atsR:C123SC187S) strains exposed to STR and H₂O₂ for 30 min. qRT-PCR assay was performed in WT(pXMJ19-atsR) strains in the absence of agents. The mRNA levels were presented relative to the value obtained from WT(pXMJ19) strains without treatment. Relative transcript levels of WT(pXMJ19) strains without stress treatment were set at a value of 1.0. Data show the averages of three independent experiments, and error bars indicated the SDs from three independent experiments. ***P ≤ 0.001 and *P ≤ 0.05 for the indicated strains versus WT strain without stress treatment. C, D Fold change of the transcription level was calculated according to the data from A, B by the equation: the value obtained from the strains exposed to stress/the value obtained from the corresponding strains without stress. Data show the averages of three independent experiments, and error bars indicated the SDs from three independent experiments. ***P ≤ 0.001 and *P ≤ 0.05 for the indicated strains versus WT strain without stress treatment. E, F The protein levels of NCgl0887 in C. glutamicum corresponding strains in the presence or absence of STR and H₂O₂. Lysates from stationary phase bacteria exposed to STR and H₂O₂ for 2 h were resolved by SDS-PAGE, and NCgl0887 was detected by immunoblotting using specific anti-NCgl0887 antibody. For the pellet fraction, cytosolic RNA polymerase α (α-RNAP) was used as a loading control. Similar results were obtained in three independent experiments, and data shown were from one representative experiment done in triplicate. G, H Relative quantified data for protein levels by Image Lab. Quantified protein expression of western blots in E, F. Densities of proteins were all justified with α-RNAP. Relative density ratios of WT without stress were set at a value of 1.0. Data show the averages of three independent experiments, and error bars indicated the SDs from three independent experiments. ***P ≤ 0.001; *P ≤  0.05