Legionella-Polymorphonuclear Leukocytes Interaction - Project Summary
Polymorphonuclear leukocytes (PMNs)/Neutrophils account for 50-60% of peripheral blood leukocytes in
humans, and are an essential part of the innate host defense against microbial infections. Quiescent PMNs
respond to high levels of glucose in vitro through a pro-inflammatory response mediated by up-regulation of
aerobic glycolysis. Importantly, PMNs respond to hyperglycemia in diabetic patients by releasing pro-
inflammatory cytokines. PMNs restrict intracellular growth of Legionella pneumophila (Lp), whose ability
to cause pneumonia is dependent on pathogen proliferation within alveolar macrophages. The Dot/Icm type
IV translocation system of Lp, which functions as a molecular syringe, injects protein effectors into the
macrophage cytosol to evade lysosomal fusion and macrophage restriction of pathogen proliferation. Despite
the Dot/Icm-mediated translocation of effectors into the PMNs cytosol, the cells restrict Lp proliferation. The
two main powerful antimicrobial machineries of PMNs to control microbial infections are the generation of
reactive oxygen species (ROS) and the antimicrobial agents within the azurophilic and specific granules. It
is not known whether any of the two major PMNs antimicrobial machineries are involved in Lp restriction.
The PMNs mount a pro-inflammatory response to Lp in vivo, but the mechanism is not known. In contrast to
macrophages, our preliminary data show that PMNs rapidly degrade the WT strain of Lp, while the Dot/Icm
translocation-defective mutant survives for at least 4h. We discovered that one of the Dot/Icm-translocated
effectors of Lp into the cytosol of human PMNs is a Legionella amylase (LamA), which degrades the PMNs
glycogen within 30-60 min of infection. Interestingly, PMNs fail to degrade the lamA-deficient mutant, which
does not degrade PMNs glycogen, similar to the Dot/Icm translocation-defective mutant. We will test the
hypothesis that human PMNs respond to the effect of the translocated LamA effector by engaging their
antimicrobial machineries in response to the cytosolic hyper-glucose generated by the LamA-mediated
degradation of glycogen. To test the hypothesis, our specific aims are: Specific Aim I: Engagement of the
PMNs antimicrobial machineries in response to the effect of LamA; and Specific Aim II: Pro-inflammatory
activation of PMNs in response to cytosolic hyper-glucose generated through glycogen degradation by LamA.
Upon completion of the proposed studies, we will learn the mechanism of the innate immune response of
human PMNs to Lp in response to abnormally high levels of cytosolic glucose. Importantly, our studies have
broad significance as they will be relevant to mechanisms of innate immunity under diabetic conditions where
dysregulated inflammation is a common persistent occurrence. Our proposed studies are novel as well as
mechanistic, and would generate a new paradigm in our knowledge of PMNs innate immune responses to
bacterial pathogens.
