Salmonella cause 1.4 million cases of gastroenteritis and enteric fever per year in the US and lead all other foodborne bacterial pathogens as a cause of death. The most serious disease results from S. typhimurium survival in phagocytes, which normally kill bacteria by producing a variety of antimicrobials including superoxide (O2-). The mechanism by which phagocytic 02- damages bacterial cells is completely unknown. S. typhimurium strain 14028 possesses two periplasmic superoxide dismutases. SodCI, encoded on the Gifsy-2 phage, contributes significantly to virulence by protecting against phagocytic 02-. The chromosomally encoded SodCII, which we have shown is expressed during infection, does not contribute to survival, even in the absence of SodCI. The two enzymes behave similarly in vitro with the notable exception that SodCI is not released by osmotic shock, a phenomenon we term """"""""tethering."""""""" This is a novel property for a periplasmic protein. We hypothesize that SodCII is produced but is non-functional, probably due to proteolytic degradation in the macrophage phagosome, and that the physical or spatial association of SodCI with some periplasmic component accounts for its preferential role in virulence.
The specific aims of this proposal are to: 1. Determine the fate of SodCI and SodCII in the phagosome. We will simultaneously monitor the production of both SodCI and SodCII protein during infection in an animal as well as in tissue culture macrophages. 2. Determine the structural and functional characteristics of SodCI that allow it to protect against phagocytic superoxide. We will exploit the differential activity of SodCI and SodCII. Hybrid proteins will be constructed and characterized. SodCs from other pathogenic bacteria and specific site directed mutants will also be tested for the ability to complement SodCI. The ability to combat phagocytic superoxide will be correlated with other characteristics including release by osmotic shock. 3. Determine if tethering to the periplasm is required for function in vivo and understand the biochemical nature of tethering. A genetic screen for mutants that are no longer tethered as well as a biochemical identification of what SodCI interacts with in the periplasm will lead to an understanding of tethering and its role in SodCI function. This research addresses a fundamental issue in innate immunity and has implications for combating a variety of important pathogens.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI063230-05
Application #
7531800
Study Section
Bacteriology and Mycology Subcommittee 2 (BM)
Program Officer
Alexander, William A
Project Start
2004-12-01
Project End
2010-11-30
Budget Start
2008-12-01
Budget End
2010-11-30
Support Year
5
Fiscal Year
2009
Total Cost
$338,449
Indirect Cost
Name
University of Illinois Urbana-Champaign
Department
Microbiology/Immun/Virology
Type
Schools of Arts and Sciences
DUNS #
041544081
City
Champaign
State
IL
Country
United States
Zip Code
61820
Golubeva, Yekaterina A; Ellermeier, Jeremy R; Cott Chubiz, Jessica E et al. (2016) Intestinal Long-Chain Fatty Acids Act as a Direct Signal To Modulate Expression of the Salmonella Pathogenicity Island 1 Type III Secretion System. MBio 7:e02170-15
Craig, Maureen; Sadik, Adam Y; Golubeva, Yekaterina A et al. (2013) Twin-arginine translocation system (tat) mutants of Salmonella are attenuated due to envelope defects, not respiratory defects. Mol Microbiol 89:887-902
Golubeva, Yekaterina A; Sadik, Adam Y; Ellermeier, Jeremy R et al. (2012) Integrating global regulatory input into the Salmonella pathogenicity island 1 type III secretion system. Genetics 190:79-90
Slauch, James M (2011) How does the oxidative burst of macrophages kill bacteria? Still an open question. Mol Microbiol 80:580-3
Rushing, Marcus D; Slauch, James M (2011) Either periplasmic tethering or protease resistance is sufficient to allow a SodC to protect Salmonella enterica serovar Typhimurium from phagocytic superoxide. Mol Microbiol 82:952-63
Kim, Byoungkwan; Richards, Susan M; Gunn, John S et al. (2010) Protecting against antimicrobial effectors in the phagosome allows SodCII to contribute to virulence in Salmonella enterica serovar Typhimurium. J Bacteriol 192:2140-9
Saini, Supreet; Slauch, James M; Aldridge, Phillip D et al. (2010) Role of cross talk in regulating the dynamic expression of the flagellar Salmonella pathogenicity island 1 and type 1 fimbrial genes. J Bacteriol 192:5767-77
Saini, Supreet; Ellermeier, Jeremy R; Slauch, James M et al. (2010) Correction: The role of coupled positive feedback in the expression of the SPI1 type three secretion system in Salmonella. PLoS Pathog 6:
Saini, Supreet; Ellermeier, Jeremy R; Slauch, James M et al. (2010) The role of coupled positive feedback in the expression of the SPI1 type three secretion system in Salmonella. PLoS Pathog 6:e1001025
Chubiz, Jessica E Cott; Golubeva, Yekaterina A; Lin, Dongxia et al. (2010) FliZ regulates expression of the Salmonella pathogenicity island 1 invasion locus by controlling HilD protein activity in Salmonella enterica serovar typhimurium. J Bacteriol 192:6261-70

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