GIT1 Regulates Spine Morphogenesis and Synapse Formation

描述（由申请人提供）：突触形成和可塑性是正常认知功能（如学习和记忆）的基础。以往的研究指出，在突触形成和可塑性的调节肌动蛋白的动力学和粘附的关键作用。这些过程可以通过GIT 1结合在一起，GIT 1是一种最近表征的分子，位于海马神经元的突触前和突触后末端。GIT1作为一个衔接子，结合Rac效应器和调节器和其他突触分子。我们的初步工作已经导致了工作假设，本提案的目的是使用分离的海马神经元和切片制备物的培养物来严格测试这些假设，如具体目标中所述。具体目标我将测试的假设，GIT 1可以通过靶向Rac突触通过组装的信号模块组成的Rac，PIX，和PAK，定位在树突棘调节突触可塑性。将使用FRET生物传感器以及我们正在开发的基于新定位的测定来测定活性Rac。将使用我们的GIT、PIX和PAK突变体文库确定GIT 1的功能。在具体目标II中，我们将测试整合素信号在突触中激活Rac的假设。该方法是确定是否整合素信号的关键调节因子，如FAK，Src和MAP激酶，调节突触可塑性。在特定目标III中，我们将使用来自海马的切片培养物将我们的观察扩展到体内类似的环境。这些研究将增强我们对突触形成和可塑性基础的分子因素的理解。

DESCRIPTION (provided by applicant): 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.