Legionella pneumophila is an intracellular pathogen that evades endocytic fusion and hijacks ER-to-golgi vesicle traffic to remodel its phagosome into an ER-derived compartment permissive for intracellular replication. The Dot/lcm type IV secretion system of L. pneumophila is essential for both modulation of phagosome biogenesis and modulation of the macrophage apoptotic pathways through robust early and novel activation of caspase-3, independent of the intrinsic and extrinsic pathways of apoptosis. The Dot/lcm- mediated activation of caspase-3 results in cleavage of Rabaptin-5, which is an effector of the early endosome regulator Rab5. Inhibition of caspase-3 using caspase-3 peptide inhibitors blocks intracellular replication of L. pneumophila and results in trafficking of the organism to phagolysosomes, similar to dot/icm mutants. Our hypothesis is: Caspase-3-meditaed cleavage of Rabaptin-5 is central to the arrested biogenesis of the Legionella-containing phagosome (LCP) and to intracellular replication. To test this hypothesis, our specific aims are to examine the following:
Specific aim I. Role of caspase-3 activation in biogenesis of the LCP;
Specific aim II : Role of caspase-3-mediated cleavage of Rabaptin-5 in biogenesis of the LCP;
Specific aim III. Identification of the L. pneumophila effector involved in caspase-3 activation. Significance: Our proposed studies are the crux of our understanding of the molecular and cellular mechanisms by which this pathogen commandeer the host cell, and will she light on the cellular mechanism by which this pathogen engages a cross talk between the apoptotic pathways and vesicle traffic in the host cell to remodel it to its liking. The bacterial effector involved in this process is a potential target for therapy and prevention of the disease.
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