The African trypanosome, Trypanosoma brucei, is a protozoan parasite that causes fatal sleeping sickness in humans and related debilitating diseases in animals. Existing treatments for these diseases are outdated, toxic and hard to administer, highlighting the need for research that opens up avenues to identify new therapeutic targets. T. brucei is capable of infecting a diverse range of mammalian hosts and is transmitted by a tsetse fly vector. In order to infect such diverse hosts, T. brucei must be able to perceive host-specific signals and respond to meet the demands presented by each host environment. These functions are largely provided by parasite cell surface proteins that form the host-parasite interface. However, to date, only a handful of T. brucei surface proteins have been identified and characterized, leaving large gaps in our understanding of host-pathogen interactions. The single flagellum of T. brucei is important for motility and is postulated to provide important signaling functions, as has been demonstrated for flagella in other eukaryotes. The flagellar membrane is proposed to be a key host-parasite interface that interacts directly with host tissues and directs signaling events important for infection and pathogenesis. By developing a novel strategy for isolation of intact membrane-enclosed flagella, combined with surface biotinylation and shotgun proteomics, we have conducted the first proteomic analysis of the flagellar membrane from bloodstream form T. brucei. Preliminary analysis of the flagellum from procyclic form parasites identified different adenylate cyclases in each life cycle stage. The function of ACs is unknown, but they are suggested to function in signaling events important for host infection. We will test the hypothesis that ACs upregulated in bloodstream form parasites mediate host-parasite interactions that affect the outcome of mammalian infection and pathogenesis. Additionally, since the flagellum is an important site of signaling and host-pathogen interaction, we will define mechanisms for targeting these receptor proteins to the flagellar membrane. Together, the proposed experiments will increase understanding of the role of trypanosome surface proteins during infection and advance efforts to understand and exploit the flagellum as a key host-parasite interface.