Project Summary The black legged tick, Ixodes scapularis, vectors several bacterial and viral pathogens, and is the major vector of Lyme disease in North America. Strategies that target this vector rather than individual pathogens stand to protect against several diseases. Thus, there is an urgent need to understand unique aspects of the tick?s biochemistry and physiology so that new targets can be identified for species-specific management strategies. Female ticks increase in size by approximately 100-fold during a several day long feeding period before dropping off the host to lay eggs. This phenomenon is barely understood at the biochemical or molecular level. A coordinated response modulating enzyme activity and structural protein production/ trafficking accompanies and enables the rapid expansion of a feeding adult female. As our first Aim, we will compare the proteomic profiles of the endocuticle (epithelial cell monolayer), which directs cuticular structure and remodeling, of unfed, ?slow phase? feeding, and ?rapid phase? feeding female ticks. We choose to work at the proteomic rather than transcriptomic level because protein levels often do not correlate with their cognate mRNAs, and protein levels provide a clearer idea of the biochemical response. This effort will produce the first comprehensive proteomic study of the cuticle of any tick. Our second Aim is to clarify the composition of the epicuticle ? the thin waxy outer layer that protects against desiccation. We challenge the paradigm that the epicuticle contains hydro- carbons. The literature notes instances of even-chain length n-alkanes and alkanes with unusual methyl branching patterns that contradict current understanding of biological hydrocarbon biosynthesis. We hypothesize that the epicuticle is dominated by dietary lipids transported directly from the midgut, and that ticks do not synthesize hydrocarbons. We will apply a multi-approach strategy incorporating proven as well as new survey methods to characterize epicuticle composition. Our work will clarify our understanding of how the tick cuticle is structured and remodeled during feeding, and correct our understanding of cuticle function. We will increase the range of molecular targets and provide new information that will accurately guide development of future tick management strategies.