Mechanistic evaluation of resistance to sulfite toxicity in Salmonella

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PROJECT SUMMARY Sulfites are reactive sulfur species with ubiquitous distribution in all life forms. They are byproducts of sulfated amino acid metabolism and have been used for decades to limit bacterial growth in food products. Because sulfites can react with proteins, DNA, and lipids to disrupt cellular processes, detoxification of sulfites is essential for cellular viability. Interestingly, sulfites are produced by activated neutrophils in response to bacterial infection, indicating they may play an important role in host-pathogen interactions. The purpose of this work is to establish how sulfites impact the outcome of bacterial infection, using Salmonella enterica as a model pathogen. Salmonella enterica is a leading cause of bacterial foodborne gastroenteritis and is the leading cause of death from foodborne disease in the United States. In our published work, we characterized the role of a transcriptional regulator, YeiE, in Salmonella pathogenesis. We demonstrated that yeiE is critical for Salmonella to colonize the intestine because it regulates production of flagella, which are a key virulence factor required for enteric salmonellosis. Recent work in Cronobacter sakazakii demonstrated that YeiE senses excess sulfite to regulate expression of a sulfite reductase and is needed to survive neutrophil killing. YeiE homologs are present in many bacterial pathogens, suggesting an important role for sulfite sensing by YeiE in host-pathogen interactions. We hypothesize that YeiE regulates the sulfite stress response to allow Salmonella to withstand innate immune control. In Aim 1, we will determine the role of sulfite reduction in salmonellosis. We will establish the impact of YeiE-regulated sulfite reduction in murine infection models of gastroenteritis and sepsis. Since activated neutrophils produce sulfite and neutrophilic inflammation is a key to host control of Salmonella, it is important to understand how YeiE mediates survival in the face of host-derived sulfite stress. In Aim 2, we will establish the mechanism by which YeiE regulates resistance to sulfite toxicity. We will establish the genes regulated by sulfite-bound YeiE using chromatin immunoprecipitation-sequencing and will establish the genes needed to withstand sulfite stress using a high-density transposon mutant library. Combined analysis of these two unbiased approaches will allow us to establish how YeiE coordinates the bacterial response to sulfite stress. YeiE homologs are distributed amongst many diverse bacterial pathogens, so our work will have application to a broad range of host-pathogen interactions. Successful completion of this work will demonstrate how bacteria resist sulfite stress in vivo and will lead to future work to investigate the role of sulfite as a cross- kingdom signaling molecule and its impact on the immune response to bacterial infections.