ROLE OF YAPS IN Y. PESTIS PATHOGENESIS

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Yersinia pestis is a Gram-negative bacterial human pathogen that is the causative agent of plague. Historically Y. pestis has been responsible for significant human morbidity and mortality. However, Y. pestis is typically a pathogen of rodents, and is unique among the Enterobacteriaceae in using a blood-sucking insect (flea) for transmission. Y. pestis infection in humans is an acute febrile disease that can have a number of different presentations depending upon the route of inoculation. Although environmental outbreaks of plague in the developed world largely have been controlled, the possible use of this pathogen as an agent of bioterrorism, as well as the re-emergence of plague in several countries, necessitates a more detailed understanding of the pathogenesis of Y. pestis to facilitate the identification of targets for vaccine development and treatment. No vaccine is currently available. A group of proteins that have a high likelihood of contributing to Y. pestis pathogenesis are the autotransporter proteins (ATs). AT proteins consist of three basic domains: a N-terminal signal sequence, a ¿passenger domain¿ and finally a β-domain at the C-terminus which facilitates translocation of the passenger domain across the outer membrane. The passenger domain provides the functional activity and new activities are being attributed to these proteins as more of them are studied in detail. While the β-domains are relatively conserved between ATs the passenger domains can vary significantly. In silico analyses have identified ten AT proteins of Y. pestis (designated yaps/Yaps). The Yaps are of interest to study for a number of reasons. First, while many putative AT proteins have been identified by in silico analyses, relatively few have been studied experimentally and even fewer have been studied in detail. With Y. pestis we have the advantage of being able to pair sophisticated molecular genetic manipulations with an animal model that is actually a natural host for this pathogen. Second, many of the Yaps by sequence analysis are relatively divergent when evolutionary trees of ATs are constructed and thus, the likelihood of identifying new functions by studying this group of ATs is high. Third, analysis of the Yaps may help identify a useful therapeutic target or candidate for inclusion in upcoming Y. pestis vaccine cocktails. We have preliminary data indicating that all ten of the yaps are expressed not only during laboratory conditions but also during infection. We also have constructed deletion mutants in seven yaps in a fully virulent Y. pestis strain and have begun testing the effect of these mutations on virulence. Four yap mutants clearly have phenotypes in a bubonic plague model of infection; these results are consistent with our hypothesis that the yaps play a role in pathogenesis. Our long term goal is to understand at a molecular level the role(s) of the individual Yaps in disease and the Y. pestis life cycle. Thus, we propose the following specific aims: (1) Analysis of the localization and expression of Yaps in Y. pestis; (2) Analysis of the role of Yaps in virulence; (3) Analysis of potential functions of the Yaps.