The long term goal of this research is to test the hypothesis that voltage-gated Na+ channel (Nav1) beta subunits are cell adhesion molecules (CAMs) that communicate between extra- and intra-cellular signaling molecules and cytoskeletal proteins. beta1 subunits are multifunctional molecules that participate in modulation of Nav1 kinetics, in extracellular matrix interactions resulting in cellular migration, in homophilic cell adhesion resulting in cytoskeletal recruitment, in heterophilic cell adhesive interactions resulting in stabilization of Nav1 complexes at the cell surface, and in neurite outgrowth from cerebellar granule neurons. beta1 can be tyrosine phosphorylated, resulting in its differential subcellular localization in cardiac myocytes. Finally, beta1 subunits contribute to the regulation of neurpnal excitability as evidenced by the effects of beta1C121W in GEFS+1 epilepsy and the severe epileptic phenotype observed in beta1 (-/-) mice. This laboratory was the first to characterize the beta subunits as CAMs and, in doing so, initiated an entirely new field in Nav1 biology. It is proposed that Nav-beta1 subunits, as CAMs, not only play important roles in cell-cell and cell- matrix adhesion, but also modulate intracellular signaling in neurons as a result of these interactions. Furthermore, it is proposed that tyrosine phosphoryation of beta1 regulates its differential subcellular targeting to differential subcellular domains in neurons and thus determines the availability of beta1 to associate with cytoskeletal and signaling molecules in those domains. The following specific aims are designed to test this hypothesis: 1. To investigate the mechanism of beta1- mediated neurite outgrowth from cerebellar granule neurons. 2. To determine the domains of beta1 that are required to mediate neurite outgrowth and to investigate whether deletion of the beta1 gene results in differences in neurite outgrowth in vivo by investigating the migration and neurite extension of corticospinal axons and cerebellar granule cells in beta1-/- mice. 3. To test the hypothesis that beta1 contains a tyrosine-based targeting domain that includes beta1Y181 and that phosphorylation of this residue results in differential localization of pYbeta1 to ankyrin-independent subcellular domains in neurons. The overall goal of this research is to investigate the novel idea that Nav1 beta subunits are not only channel modulators but also act to initiate signal transduction cascades as a result of cell adhesive interactions. A beta1 mutation that causes epilepsy in humans, beta1C121W, not only disrupts beta1-mediated channel modulation but also disrupts beta1-mediated cell adhesion. Thus, as genetic mutations that affect L1-CAM-mediated cell adhesion result in severe neurological defects such as hydrocephaly and mental retardation, it is proposed that human mutations that result in disruption of Nav1 beta subunit-mediated cell adhesion may also result in neurological disease, including epilepsy.
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