B-287. Identification and Characterization of Four Autotransporter Proteins in Yersinia pestis

D. G. Cotter1,2, M. B. Lawrenz2, J. D. Lenz2, W. Lathem2, W. E. Goldman2, V. L. Miller2;
1Univ. of Nevada, Las Vegas, NV, 2Washington Univ. Sch. of Med., St. Louis, MO.

In relation to other secretion systems, the autotransporter, or type V, secretion system is unique in its simplicity. Autotransporters contain within their primary amino acid sequence all the information necessary for their transport to the outer membrane and are characterized by an overall unifying structure: an N-terminal signal sequence, a variable passenger domain, and a C-terminal β-domain that mediates passage of the mature passenger domain across the outer membrane to the cell surface. The passenger domains of autotransporters encode a diverse range of functions, many of which have been implicated in virulence. In silico analyses of the genome of the plague pathogen Yersinia pestis revealed the presence of 11 putative autotransporter proteins_ designated the Yaps (Yersinia autotransporter proteins). These genes were shown to be expressed during mammalian infection by RT-PCR, suggesting a role in virulence. We used a tetracycline inducible system to express epitope tagged versions of the Yaps in Escherichia coli DH5α. Surface proteolysis, cell fractionation, and Western blot analysis were performed to determine the localization of YapA, YapF, YapM, and YapG. Based on studies of other autotransporters we predicted that these Yaps would be in the outer membrane, or released into the supernatant. All these Yaps localized to the outer membrane and furthermore, YapG and YapA appear to be processed and released into the supernatant. In the murine model for bubonic plague, mutations in YapG and YapF resulted in altered rates of dissemination, indicating that these autotransporters may encode functions important for pathogenesis.