Protection from Oxidative Stress Relies Mainly on Derepression of OxyR-Dependent KatB and Dps in Shewanella oneidensis

Abstract

Shewanella thrives in redox-stratified environments where accumulation of H 2 O 2 becomes inevitable because of the chemical oxidation of reduced metals, sulfur species, or organic molecules. As a research model, the representative species Shewanella oneidensis has been extensively studied for its response to various stresses. However, little progress has been made toward an understanding of the physiological and genetic responses of this bacterium to oxidative stress, which is critically relevant to its application as a dissimilatory metal-reducing bacterium. In this study, we systematically investigated the mechanism underlying the response to H 2 O 2 at cellular, genomic, and molecular levels. Using transcriptional profiling, we found that S. oneidensis is hypersensitive to H 2 O 2 in comparison with Escherichia coli, and well-conserved defense genes such as ahpCF, katB, katG, and dps appear to form the first line of defense, whereas iron-sulfur-protecting proteins may not play a significant role. Subsequent identification and characterization of an analogue of the E. coli oxyR gene revealed that S. oneidensis OxyR is the master regulator that mediates the bacterial response to H 2 O 2 -induced oxidative stress by directly repressing or activating the defense genes. The sensitivity of S. oneidensis to H 2 O 2 is likely attributable to the lack of an inducible manganese import mechanism during stress. To cope with stress, major strategies that S. oneidensis adopts include rapid removal of the oxidant and restriction of intracellular iron concentrations, both of which are achieved predominantly by derepression of the katB and dps genes.

Defining the binding determinants of Shewanella oneidensis OxyR: Implications for the link between the contracted OxyR regulon and adaptation

“…In SoOxyR, the counterparts of these two cysteines are Cys-203 and Cys-212 respectively; mutation of either residue to serine locks SoOxyR into the reduced form (19,24). In addition, SoOxyR L197P mutant is found to be locked into the oxidized state (25) (18,19). Because the promoter is slightly leaky (26,27) Fig.…”

“…Recombinant His-tagged OxyR proteins exist in reduced and oxidized forms -S. oneidensis OxyR, as a H 2 O 2 -responsive transcriptional regulator, empowers cells to confront oxidative stress by regulating gene expression through reversible formation of intermolecular disulfide bond (19). In order to characterize the biochemical properties of this regulator in more detail, we expressed recombinant SoOxyR with hexahistidine(his 6 )-tag at the N-terminal in E. coli.…”

“…Second, SoOxyR is also able to respond to organic peroxides (OP), functionally intertwined with OhrR, a regulator specific for OP (23). Third, SoOxyR is unable to complement the E. coli oxyR mutation and vice versa although they share a comparable sequence similarity as mentioned above (11,13,19). Hence, SoOxyR apparently mediates cellular response to oxidative stress with unprecedented mechanisms.…”

“…Nevertheless, three unexpected phenotypes resulting from the SooxyR disruption were observed (11). First, although the SooxyR mutant under non-stress conditions degrades H 2 O 2 more rapidly, it still carries defects in both viability (plating defect) due to increased sensitivity to H 2 O 2 and growth (18,19,20), phenotypes reported in bacteria with OxyR being a positive regulator, such as E. coli (21; 22). Second, SoOxyR is also able to respond to organic peroxides (OP), functionally intertwined with OhrR, a regulator specific for OP (23).…”

“…Because of the potential application in bioremediation, biogeochemical circulation of minerals and bioelectricity, these bacteria have been intensively studied, especially the model species S. oneidensis (18). The S. oneidensis OxyR (SoOxyR) resembles those of Neisseria in that it represses expression of catalase gene katB under non-stress conditions but functions as an activator for all of its target genes when cells are challenged by H 2 O 2 (19). Nevertheless, three unexpected phenotypes resulting from the SooxyR disruption were observed (11).…”