The work in my laboratory is directed toward investigating the molecular mechanisms of bacterial uptake and intravacuolar growth in host cells. The investigation of the Yersinia pseudotuberculosis invasin protein has been the primary focus of our studies on uptake, and analyses of intravacuolar growth have been performed using Legionella pneumophila. Genetic analysis of these organisms is relatively facile, and each promotes cellular events that are observed with many other pathogens. The most exciting recent development is our identification of effector proteins that are translocated into target cells by L. pneumophila and the characterization of signaling pathway for uptake of Yersinia.

Intracellular growth of Legionella pneumophila
L. pneumophila avoids phagocyte killing by growing within a replication vacuole that initially bypasses the lysosomal network. The bacterium is internalized into a membrane-bound compartment that recruits endoplasmic reticulum-derived vesicles as a prelude to the wholesale docking of rough endoplasmic reticulum. Formation of this replication vacuole requires the products of 26 dot/icm genes, which encode proteins that assemble into an apparatus that translocates proteins into target host cells. We have devoted recent work to identifying proteins that are translocated via this apparatus. These translocated substrates appear to associate with cytoplasmic surface of the membranous compartment surrounding the bacteria.

Cellular uptake of Yersinia pseudotuberculosis
Yersinia pseudotuberculosis is internalized by M cells overlying the intestinal Peyer’s patches Shortly after this event, bacteria are found exclusively extracellularly due to the production of Yops, translocated bacterial proteins that antagonize phagocytosis. We have been interested in internalization, the earliest stage of the infection process. Our work demonstrated that the primary bacterial-encoded factor that allows uptake both into M cells and into cultured cells is the outer membrane protein invasin, which binds multiple β1 integrin receptors. We identified residues in the protein that are involved in integrin recognition and modulation of uptake efficiency. In addition we have been investigating the cellular route that allows bacteria to translocate across the intestine. Our recent work has demonstrated that a key signaling molecule downstream from the integrin is the small GTPase, Rac1, which directs actin rearrangements in response to bacterial binding. Much of the focus of our research is directed toward identifying the proteins that are activated in response to signals sent from Rac1.

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