Hypochlorous Acid Stress Response in Bacteria.

Abstract

To kill invading bacterial pathogens, the antimicrobial hypochlorous acid (HOCl) is produced in the cells of the innate immune system. HOCl is also generated at mucosal barrier epithelia, suggesting the importance of HOCl-induced responses for bacterial pathogenesis and colonization. Studies have demonstrated that the conserved E. coli Hsp33 chaperone has a major role in protecting bacteria against oxidative protein unfolding, a stress condition mediated by HOCl. To understand the function of Hsp33 in Vibrio cholerae, a causative agent of cholera, I characterized and discovered that the V. cholerae ΔHsp33 mutant has temperature sensitive (ts) phenotype for growth when cultivated under aerobic conditions. Studies revealed that overexpression of the E. coli elongation factor EF-Tu is necessary and sufficient to rescue the ts growth defect and the severe HOCl-sensitivity in V. cholerae ΔHsp33 mutant. Mechanistic studies revealed that Hsp33 protects Vibrio EF-Tu both from degradation at normal growth temperatures, and against aggregation under stress-conditions. These results suggest that Hsp33 protects bacteria against HOCl-mediated cellular death by preventing EF-Tu, an essential component for protein biosynthesis, from oxidative protein degradation. To investigate the transcriptional changes in bacteria in response to HOCl treatment, Affymetrix microarray studies were performed on E. coli cells. Results confirmed that HOCl treatment leads to the accumulation of unfolded and aggregated proteins as the heatshock regulon were up-regulated. Protein unfolding is the likely reason for increased intracellular level of free metals, which require the up-regulation of efflux systems. I identified the up-regulation of the HOCl-specific transcriptional repressor NemR, and whose gene regulation gloA facilitates methylglyoxal detoxification in HOCl-treated bacteria. The beneficial role of methylglyoxal formation appears to be the replenishment of phosphates needed for ATP regeneration and other biological building blocks. This seems to be important as HOCl treated cells were found to accumulate large quantities of polyphosphate crystals, which play a protective role against macromolecular damage in cells. GloA-mediated methylglyoxal detoxification along with polyphosphate production, therefore appear to play major roles in cellular survival against HOCl. Results of this study will help to reveal the molecular action of HOCl and elucidate the bacterium’s strategy to counteract HOCl-stress in host defense.