This lack of sensitivity to multiple antibiotics suggests that the sigE mutation does not lead to an overall increase in the permeability of the outer membrane, which would allow more of the antibiotic to enter the cell. These
results show that SigE is important for survival in response to specific types of damage to the cell envelope, such as disruption of cellular membranes caused by SDS/EDTA and interference with synthesis of the peptidoglycan layer caused by ampicillin and mecillinam. We next asked if sigE is important for survival following a shift to high temperature, which perturbs both the cell envelope and cytoplasm. RB50 and RB50ΔsigE were grown at 37°C to an OD600 of 0.4, then shifted to 50°C, a Eltanexor nmr lethal temperature for B. bronchiseptica. Cell viability, assessed by CFU/ml, was measured after the shift to 50°C. Survival of the RB50ΔsigE strain Bafilomycin A1 supplier was lower than
that of RB50 (Figure 2C). In attempting to complement this phenotype, we found that plasmid-encoded sigE did not restore survival during heat shock (data not shown), although it did complement other phenotypes, as described below. Similar variability in complementation of a σE mutant by a plasmid-encoded rpoE gene has been seen in other bacteria [29, 36, 40, 41]. Work from Burkholderia cenocepacia CDK inhibitor drugs showed that expressing σE from a plasmid actually increased Axenfeld syndrome sensitivity to heat stress [36]. In S. Typhimurium, an rpoE mutant was sensitive to paraquat and did not survive in stationary phase under anaerobic conditions. Expression of rpoE from a plasmid partially complemented the former phenotype, but not the latter [29]. Because the anti-sigma factor
that regulates σE activity was not included in any of these instances, it is likely that proper regulation of SigE activity is required for optimal response to particular stresses, not merely excess SigE activity, complicating complementation experiments. Another aspect of the classical heat shock response is thermotolerance. When bacteria are exposed to an elevated but nonlethal temperature, heat shock responses are induced, resulting in increased production of chaperones and proteases that refold or degrade unfolded proteins [42]. Consequently, the cells are preloaded with protective factors and exhibit increased survival following a subsequent shift to a lethal temperature [42]. To investigate the role of SigE in this phenomenon, RB50 and RB50ΔsigE were grown to an OD600 of 0.1 at 37°C, shifted to 40°C for 90 min, then shifted to 50°C. RB50 cultures incubated at 40°C before 50°C survived better at all time points than those directly shifted from 37°C to 50°C.