Project Summary Endocytosis plays a crucial role in supporting the health of the human brain. Functionally, synaptic vesicle endocytosis allows neurons to sustain synaptic transmission without exhausting the supply of synaptic vesicles. Structurally, endocytosis supports the maintenance of synapses and neural circuits. As a result, defective synaptic vesicle endocytosis creates deficits in neurotransmission that underlie a wide spectrum of neurological diseases and psychiatric disorders. The long-term goal of this study is to determine how endocytic proteins act in concert to support diverse routes of endocytosis at synapses. Work from several laboratories including my own has found that the curvature-sensing protein endophilin plays a critical role in synaptic vesicle endocytosis. In this proposal, we will examine the hypothesis that curvature-sensing mechanisms guide endocytic proteins to perform their function in various routes of endocytosis. We propose three Specific Aims. 1) We will determine the role of curvature-sensing motifs in vivo. We focus on the curvature-sensing amphipathic helix of endophilin as it is essential for synaptic vesicle endocytosis. 2) We will study how curvature signals are received by the downstream protein synaptojanin to support synaptic vesicle endocytosis, and to prevent age-dependent decay of synaptic transmission. 3) We will determine the mechanism of endophilin-independent endocytosis, an area that lacks molecular information. Through an unbiased genetic screen, we have identified another curvature-sensing protein that acts in a parallel pathway with endophilin. Results from these studies are expected to to push boundaries of current knowledge of synaptic biology and to lead toward solutions for neurological disorders.