? Chemical synapses are ultrastructurally distinct subcellular entities that mediate the information flow from neurons to their targets. In the presynaptic terminals, synaptic vesicles are arranged orderly around the electron-dense active zones. Recent studies have revealed multiple pathways controlling presynaptic differentiation. A focus of this lab is to take a genetic approach in the nematode C. elegans to identify genes that regulate synapse formation. The rpm-1 gene (for regulator of presynaptic morphology) encodes a large evolutionarily conserved protein that contains multiple functional domains including a Ring-finger E3 ubiquitin ligase domain. Loss of function in rpm-1 results in diverse synaptic defects, ranging from failure to form stable synapses to disorganized presynaptic terminals. Through genetic screens, we have identified a MAP kinase module whose activity is inhibited by rpm-1. This MAP kinase module is composed of DLK-1, a MAP kinase kinase kinase; MKK-4, a MAP kinase kinase; and PMK-3, a p38-like MAP kinase. Loss of function in any one of the kinases suppresses the synaptic defects in rpm-1 mutants, whereas elevating the MAP kinase signaling in wild type animals causes abnormal synapses resembling those of rpm-1 mutants. DLK-1 is localized to presynaptic regions, and its abundance appears to be elevated in rpm-1 mutants. Based on these data, we hypothesize that during synapse formation RPM-1 functions to down-regulate the MAP kinase pathway, possibly by targeting DLK-1 for degradation. The main goals of this renewal application are to define the biochemical interactions between the MAP kinases (in particular DLK-1) and RPM-1, and to identify additional components of this MAP kinase pathway and other genes that may interact with RPM- 1. RPM-1 is localized to a distinct subsynaptic domain. Additional experiments are proposed to examine how its synaptic localization is regulated. The outcome of this application will advance our understanding of the regulatory network in synapse formation. Synapse integrity is detrimental to the function of the brain, hence the health of a human being. This study will contribute to the understanding of the basic mechanisms that maintain healthy synapses, and may also provide insights into the pathogenesis of synapse dysfunction. ? ?
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