Inhibitors of Type III Secretion and Translocation in Yersinia

Many gram negative bacterial pathogens, including Yersinia, Salmonella, Pseudomonas, Shigella,enteropathogenic E. coli, Chylamydia, and Burkholderia, use type III secretion systems (TTSS) to translocateeffector proteins from the bacterial cytosol into mammalian cells. Translocated effector proteins, called Yops inYersinia, subvert normal host processes to promote the survival of the pathogen, and thus TTSS play anessential role in the infectious process and virulence of these bacteria. TTSS are comprised of a base, whichspans the inner and outer membranes of the bacteria, a needle, which extends from the base to the host cell,and a translocon, which is inserted into host cell membranes and through which Yops are thought to travel toreach host cell cytoplasm. <P> In Yersinia, the needle is primarily composed of one 7kD protein, YscF, whichpolymerizes to form a long tube. The needle is thought to be a conduit for the passage of Yops from thebacteria to the translocon and is thought to conduct signals from the host cell membrane to the TTSS basewhen the pathogen comes in contact with a host cell. These signals trigger Yop translocation. A tip protein,LcrV, is found at the distal end of the needle. One function of LcrV is to position, assemble, and/or insert thetranslocon, into plasma membranes, which is essential for Yop delivery into cells. Two proteins, YopB andYopD comprise the translocon. <P> The overall goals of this proposal are to understand the molecular interactionsbetween TTSS components that are required for translocating Yops upon contact with mammalian cells. Wehave identified 3 dominant negative YscF alleles and 8 small molecules that inhibit Yop translocation. Theseunique mutants and compounds will be studied intensively as probes to understand the required molecularinteractions for Yop translocation into host cells. <P> Our working hypothesis is that our inhibitors interfere withcritical interactions between YscF, LcrV, YopB and/or YopD. Understanding the mechanism of translocationand needle assembly will support the development of new strategies that interfere with these processes andthus neutralize the bacteria's pathogenecity.