Dysregulation of the inflammatory response by Francisella tularensis

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Tularemia is a potentially fatal disease and the causative agent, Francisella tularensis (Ft), is one of few bacterial pathogens that can infect both neutrophils (polymorphonuclear leukocytes, PMNs) and macrophages. Notably, macrophages and neutrophils appear to play distinctly different roles in tularemia pathogenesis, with macrophages acting as major vehicles for bacterial growth and dissemination, and PMNs playing a central role in host tissue destruction. Neutrophils are short lived, and unlike other leukocytes are preprogrammed to undergo apoptosis 24 h after release into the circulation. Tight spatial and temporal control of this process is critical for elimination of infection and resolution of inflammation, and for this reason defects in PMN turnover exemplify a dysregulated and ineffective inflammatory response that promotes tissue destruction and disease. In keeping with this, we discovered that Ft inhibits human neutrophil apoptosis and markedly prolongs cell lifespan, and demonstrated that this is achieved via effects on the intrinsic and extrinsic apoptosis pathways, as well as changes in the neutrophil transcriptome that include significant differential expression of 365 unique genes linked to apoptosis and cell fate. Nevertheless, how cell lifespan is prolonged is only partially understood. Herein, we propose groundbreaking studies based on our discovery of neutrophil metabolic reprogramming as a new mechanism for apoptosis inhibition. Our proposed studies are supported by extensive preliminary data, and are highly innovative, as integrated manipulation of glycolysis and organelle function has not been previously documented as a mechanism for regulation of PMN lifespan during infection. Potential effects of these changes on bacterial growth and the influence of PMN metabolites on macrophage polarization will also be determined. In addition, we recently identified bacterial lipoproteins (BLPs) as active factors in Ft conditioned medium (CM) that extend PMN lifespan via a mechanism that is dependent on a common single nucleotide polymorphism (SNP) in human TLR1 (rs5743618, T1805G) that significantly influences the severity and lethality of sepsis as well as the outcomes of many infectious and inflammatory diseases, including but not limited to tuberculosis, pyelonephritis, atherosclerosis, arthritis, lupus, colitis, and cancer. Elucidating the mechanism(s) of BLP and TLR2/1-driven apoptosis inhibition is a second objective of this study. Our experimental design will also utilize drugs that specifically target HIF-1 and TLR2, mitophagy, glycolysis or other relevant signaling intermediates to identify points for therapeutic intervention that are expected to be relevant to many diseases that affect Veterans. Our specific aims are: 1) To elucidate the mechanisms and functional consequences of neutrophil metabolic reprogramming. 2) To elucidate the mechanisms of BLP and TLR2/1-mediated apoptosis inhibition and potential for theraputic intervention.