Successful intracellular pathogens have developed diverse strategies to avoid the lysosome. Several modify their phagosome into a specialized replicative vacuole, which either totally resists fusion with lysosomes or follows a distinct pattern of maturation, while others escape the phagosome and reside within the host cell cytosol. The goal of this work is to define the mechanisms by which these pathogens avoid phagosome- lysosome fusion. Preliminary data indicates that Synaptotagmin VII (Syt VII), a transmembrane calcium binding protein localized to lysosomes, may be involved in phagosome-lysosome fusion. If so, Syt VII may be a target of bacterial effectors designed to help the pathogen avoid phagosome-lysosome fusion.
The specific aims of this proposal are: (1) To characterize the role of Synaptotagmin VII in endosome-lysosome transport in mammalian cells using confocal and electron microscopy. (2) To determine whether Syt VII-dependent inhibition of phagosome- lysosome fusion will rescue the replication of Salmonella, Legionella, and Listeria strains defective for intracellular survival. (3) To identify potential bacterial effector proteins that interact with Syt VII using a yeast two-hybrid screen. The characterization of these potential effectors will be a first step in understanding the molecular mechanisms by which bacterial pathogens avoid phagosome-lysosome fusion and may reveal new targets for antibacterial therapy.
