Biologic Activities of the pertusis toxin B-pentamer

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DESCRIPTION (provided by applicant): Pertussis toxin (PTX) is a complex AB5 toxin, comprised of the enzymatically active, "A" subunit (S1), and the binding, "B" oligomer (PTX-B), composed of five subunits S2, S3, S4, and S5, found in a 1:1:2:1 ratio. The B portions of AB toxins transport the A subunit to the cytoplasm of target cells, and were not thought to participate in toxicity. However PTX-B has been shown to have activity in addition to its role in facilitating entry of S1 into the cytoplasm. PTX-B induces a spectrum of cellular responses, including immediate cell death, development of apoptosis, cellular clustering, and even cellular proliferation. In this proposal we will identify the receptors for PTX-B, we will characterize the signaling pathways that are affected by binding of PTX-B using T cells as a model system, and we will characterize its toxicity to other cells of the immune system. Specific Aim 1. Characterization of the PTX-B binding elements. PTX-B has been shown to bind to multiple receptors on cells. We will identify the essential PTX-B binding regions as a prelude to the identification of potential cell surface receptors. Specific Aim 2. Determine the mechanism by which PTX-B attenuates chemokine receptor signaling and chemotaxis. The mechanism by which PTX-B affects chemokine receptor activity remains largely unknown. We will extend our studies on the mechanism(s) of PTX-B signaling in T-cells to determine if PTX-B blocks lymphocyte migration by promoting chemokine receptor desensitization using the chemokine receptor CCR5 as a model system. Specific Aim 3. Characterize the ability of primary cells to respond to intact pertussis toxin and PTX-B. Published reports suggest that the responses to PTX-B are different between mice and humans. We propose to characterize the short term and long-term responses of human and murine leukocytes both intact pertussis toxin and PTX-B. Pertussis toxin is the major virulence factor of Bordetella pertussis, the causative agent of human whooping cough or pertussis. Whooping cough is common, endemic, and consumes a significant amount of health care resources, and is the only vaccine-preventable disease that is increasing in incidence in the United States. B. pertussis is able to infect previously infected or vaccinated individuals due to its ability to impede the development of a protective immune response, and understanding the interaction of pertussis toxin with the immune system is important for developing improved pertussis vaccines and therapeutics.