Fig. 3.

Effects of H₂O₂ on promoter occupancy and expression of Irr-regulated genes. Cultures were grown to mid-log phase in media supplemented with 2 μM FeCl₃ and 50 μM MnCl₂. At time 0, 2 mM H₂O₂ was added to the cultures and were continually aerated at 29°C. Cells were grown in 20 μM (+Fe) without H₂O₂ treatment as a control. Aliquots were harvested at indicated time points for analysis. (A) Steady-state levels of Irr and GroEL were detected by immunoblotting as described in Fig. 1. (B–G) Promoter occupancy of respective genes by Irr in cells treated with H₂O₂ were carried out as described in the Fig. 1 legend. The data are expressed as the relative starting quantities (SQ) of immunoprecipitated DNA normalized to the input DNA and are presented as average of three replicates ± the standard deviation. (H–M) Steady-state transcript levels of the respective genes obtained from cells treated with H₂O₂ were analyzed by qPCR as described in the Fig. 1 legend. The data are expressed as relative starting quantities (SQ) of respective mRNAs normalized to the housekeeping gene gapA, and presented as average of three replicates ± the standard deviation.