Synapse formation and plasticity underlie normal cognitive functions, such as learning and memory. Previous studies point to a critical role for the regulation of actin dynamics and adhesion in synapse formation and plasticity. These processes can be brought together by GIT1, a recently characterized molecule that resides in pre and postsynaptic terminals of hippocampal neurons. GIT1 serves as an adapter that binds Rac effectors and regulators and other synaptic molecules. Our preliminary work has led to working hypotheses and the objective of this proposal is to rigorously test these hypotheses as outlined in the specific aims using cultures of dissociated hippocampal neurons and slice preparations.
Specific Aim I will test the hypothesis that GIT1 can regulate synaptic plasticity by targeting Rac to synapses through the assembly of a signaling module comprised of Rac, PIX, and PAK that localizes in dendritic spines. Active Rac will be assayed using FRET biosensors as well as a new localization based assay that we are developing. The function of GIT1 will be determined using our library of GIT, PIX, and PAK mutants.
In Specific Aim II, we will test the hypothesis that integrin signaling activates Rac in synapses. The approach is to determine whether key regulators of integrin signaling, such as FAK, Src, and MAP kinase, modulate synaptic plasticity.
In Specific Aim III, we will use slice cultures from the hippocampus to extend our observation to an in vivo like environment. These studies will enhance our understanding of the molecular factors that underlie synapse formation and plasticity. ? ?
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