Role of Cryptosporidium Mucins in Host-parasite interactions

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DESCRIPTION (provided by applicant): Cryptosporidium is a ubiquitous waterborne pathogen that causes diarrheal disease worldwide. This pathogen is of significant medical importance because of its capacity to cause life-threatening diarrheal disease in immunocompromised hosts, particularly AIDS patients. To date there is no consistently effective treatment for Cryptosporidium infection, and no vaccine to prevent infection. Studies suggest that Cryptosporidium, unlike other apicomplexans, relies on mucin-like glycoproteins to attach to and invade intestinal epithelial cells. These mucin antigens are also targets of protective cellular immune responses. Despite the importance of mucin antigens to Cryptosporidium biology, few of these antigens have been investigated. The Cryptosporidium genome project has identified 31 open reading frames encoding putative mucin antigens. The studies proposed in this application will investigate 7 mucins that reside on a single locus on chromosome 2 (CpMuc1-7) and determine the role they play in host-parasite interactions. Aim #1: To characterize expression, localization, post-translational modifications, and potential multi-protein complex formation of the seven mucin-like glycoproteins of the CpMuc1-7 locus. We will quantify expression of the CpMucs during intracellular development using real-time RT-PCR, determine protein expression and localization, elucidate the glycosylation patterns of the native antigens, and determine if these antigens form multi-protein complexes. Aim #2: To determine if the CpMuc1-7 glycoproteins play a role in invasion and/or stimulation of cell mediated immune responses. We will determine if anti-CpMuc antibodies and recombinant proteins inhibit C. parvum invasion of intestinal epithelial cells, and determine if the CpMucs are activators of cellular immune responses that develop during infection in mice. In these studies, we will also explore the effect of glycosylation in host cell invasion and immune activation. Finally, employing a non-virulent T. gondii strain as an antigen delivery system, we will determine if immune responses targeted against one of the CpMucs provides protection from cryptosporidiosis. These studies will enhance our understanding of the role mucin-like glycoproteins play in the establishment and resolution of Cryptosporidium infections and greatly advance our ability to design effective treatments to combat this serious infectious threat around the world.