Elucidation of the regulation mechanisms that control the degradation of synaptic vesicle proteins represents a critical step towards our goal of understanding how synaptic function is regulated under physiologic conditions and how synaptic terminals become dysfunctional and degenerated in diseases. Synaptic vesicle proteins play a critical role in neurotransmitter release, and changes in the expression levels of synaptic vesicle proteins contribute to synaptic plasticity such as learning and memory. Furthermore, alterations in the expression levels of synaptic vesicle proteins are associated with a variety of neurodegenerative diseases and psychiatric disorders including drug addiction. Despite the importance of the regulation of synaptic vesicle protein levels in synaptic function and dysfunction, the molecular mechanisms underlying such regulation remain uncharacterized. On the other hand, the ubiquitin-proteasome proteolytic pathway has emerged as a major mechanism by which cells regulate the expression levels of specific proteins, and aberrations in the ubiquitin-proteasome pathway have been implicated in the pathogenesis of several neurodegenerative diseases. However, very little is known about neuronal protein substrates that are targeted by this pathway. The applicant hypothesizes that the ubiquitin-proteasome pathway targets synaptic vesicle proteins for degradation through specific E3 ubiquitin-protein ligases. In support of this hypothesis, our preliminary studies suggest that Siah-1 and Siah-2, a family of mammalian Seven in Absentia homologues, may act as E3 ubiquitin-protein ligases to regulate the degradation of synaptic vesicle protein synaptophysin. In this project, this hypothesis will be tested thoroughly. The specific aims are to: 1) characterize Siah-mediated ubiquitination and degradation of synaptophysin at nerve terminals; 2) investigate the molecular mechanisms by which Siah-1 and Siah-2 regulate the ubiquitin-dependent degradation of synaptophysin; 3) identify additional synaptic targets of Siah proteins; 4) determine the distribution and subcellular localization of Siah proteins in the central nervous system; and 5) examine the role of Siah-mediated protein degradation in neuronal apoptosis. These studies should advance our understanding of protein degradation at synaptic terminals, and provide a basis for the development of therapeutic strategies for treating malfunctions of the nervous system.