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Identifying Targets for Therapeutic Interventions of Salmonella Bacteria and Orthopox Viruses Using Proteomic Technology

Josh Adkins, PNNL Contact
Richard Smith, Principal Investigator

Funding Agency: National Institutes of Health, National Institute of Allergy and Infectious Disease

Closely related pathogens vary substantially in virulence. Consider distinct species of Yersinia (e.g., pestis and enterocolitica), Salmonella (e.g., typhimurium, typhi, and arizona ), and related orthopoxviruses (e.g., variola major, vaccinia, and monkeypox). Even within the same species, many mutant strains are totally avirulent in immunocompetent individuals. Although the precise genetic defect is known for many strains of decreased virulence, the consequences of even well-defined mutations on the expression of other proteins by the pathogen is unknown, particularly the major intracellular regulatory proteins. The Identifying Targets for Therapeutic Interventions of Salmonella Bacteria and Orthopox Viruses Using Proteomic Technology research team at Pacific Northwest National Laboratory (PNNL) and Oregon Health & Science University hypothesizes that a subset of the proteins differentially expressed in wild-type versus mutant strains of Salmonella is mechanistically involved in pathogenicity, and thus could be a novel therapeutic target. In a parallel series of experiments involving orthopox viruses, we hypothesize that proteins differentially expressed in wild-type monkeypox, compared to vaccinia or an avirulent mutant strain of monkeypox, contribute mechanistically to the differing virulence of these viruses in human hosts.

Salomella tryphimurium
Salmonella typhimurium is closely related to the causative agent of typhoid fever, and it infects approximately 40,000 people each year in the U.S. Protein measurements from S. typhimurium harvested under different culture conditions indicate that most proteins did not change significantly. A subset of proteins whose abundance was largely restricted to a medium mimicking host growth was studied in more detail.

These hypotheses form the rationale for a comparative proteomic study of wild type and mutant Salmonella strains grown in defined culture and in distinct cell types, and for parallel proteomic studies of defined orthopox virus strains grown in epithelial cells and macrophages. We find that comparisons of virulent and mutant or avirulent proteomes identify a number of differentially expressed proteins, many of which were not recognized to be involved in pathogenesis. Similarly, comparisons of the expressed proteins from the same strain cultured in different cell types will also reveal many differences, only a subset of which contribute to pathogenesis. However, we hypothesize that those genes that are under the control of major intracellular regulators are more likely to be required for virulence because their expression is tightly regulated in time and space. Obtaining specific information about the timing of expression, particularly in relation to the timing of intracellular replication, will provide important information regarding those gene products most essential for intracellular replication. Based on this assumption, we will use a bioinformatic approaches to select for further validation those proteins associated with avirulence that are also altered in non-productive infections.

A key advantage of performing much of this research on the PNNL campus is the solid technical foundation in advanced proteomic technologies already established within the Environmental Molecular Sciences Laboratory housed at PNNL. The research benefits from an existing large suite of cutting-edge instrumentation and data processing capabilities, as well as an established cross-disciplinary team of scientists. Additional benefits include an existing NIH collaboratory infrastructure through the Proteomics Research Resource for Integrative Biology and PNNL’s extensive support for major instrumentation and baseline operational costs.

More information about this research can be found on its corresponding NIAID Administrative Center page.

Data can be found at the PNNL Data Section of the NIAID Administrative Resource for Biodefense Proteomics Research Centers.

Related Publications

Adkins JN, HM Mottaz, AD Norbeck, JK Gustin, J Rue, TR Clauss, SO Purvine, KD Rodland, F Heffron, RD Smith. 2006. "Analysis of the Salmonella typhimurium proteome through environmental response toward infectious conditions." Molecular and Cellular Proteomics 5(8):1450-1461.

Shi L, JN Adkins, JR Coleman, AA Schepmoes, A Dohnalkova, HM Mottaz, AD Norbeck, SO Purvine, NP Manes, HS Smallwood, H Wang, J Forbes, P Gros, S Uzzau, KD Rodland, F Heffron, RD Smith, TC Squier. 2006. "Proteomic analysis of Salmonella enterica serovar typhimiurium isolated from RAW 264.7 macrophages: Identification of a novel protein that contributes to the replication of serovar typhimiurium inside macrophages."Journal of Biological Chemistry 280(39):29131-29140.

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