Signaling Mechanisms Regulating Rac-dependent Synaptic and Dendritic Development

项目总结 大脑中神经回路的形成和信息处理需要精确控制发育和 富含肌动蛋白的树突棘和它们所容纳的兴奋性突触的重塑。动态调节 介导快速兴奋性突触传递和突触的AMPA和NMDA型谷氨酸受体 可塑性分别是这种控制的一个关键方面。突触病理是许多脑部疾病的特征 包括智力残疾、自闭症、躁郁症、抑郁症和阿尔茨海默病。因此， 揭示控制脊椎/突触发育和谷氨酸受体调节的机制将 提供有关大脑功能和疾病的重要见解。Rho GTP酶是脊椎/突触的主要调节者 发展和重塑。Rac1促进脊椎/突触的形成、生长和维持，而 RhoA抑制这些过程；两者在突触可塑性中也起着关键作用。Rho的正常功能 GTP酶需要精细的时空控制，这种调控的破坏导致大量大脑 精神错乱。Rho GTP酶被鸟嘌呤核苷酸交换因子(GEF)激活，并被 GTP酶激活蛋白(GAP)。然而，令人惊讶的是，人们对这些GEF/间隙是如何形成的知之甚少 指导神经形成的时空rac1/RhoA激活模式和效应器反应 大脑中的电路。我们发现，rac1-gef Tiam1是树突、棘突和突触去突触的关键调节因子。 发育，表明它将突触受体偶联到rac1激活和肌动蛋白细胞骨架 培养的海马神经元的重塑。在上一个赠款周期中，我们有了一个令人惊讶的发现 Tiam1与rac1-GAP/RhoA-GAP BCR结合，需要这种全环基金/GAP复合体来精确调控 突触的rac1信号和兴奋性突触的形成。BCR与双相情感障碍和学习障碍有关 行为缺陷，而Tiam1的表达变化在抑郁症和唐氏综合症患者中可见。 我们假设Tiam1/BCR协同控制Rho的激活动力学和信号特异性 GTP酶，在体内是脊椎/突触正常发育所必需的，NMDAR运输/功能， 学习和情绪调节。为了测试这一点，我们建议：(1)确定Tiam1和密切相关的角色 Tiam2在体内形成脊柱/突触发育中的作用及其影响的具体途径；以及 (2)阐明Tiam1/BCR在突触可塑性、学习和心境中调控NMDARs的机制 监管。我们将使用一种多学科的方法，包括小鼠遗传学、延时活细胞和体内 双光子成像、Förster共振能量转移、电生理学、生物化学、分子 以及细胞生物学和行为分析。我们的发现将阐明控制Rho的关键机制 GTPase依赖的突触发育/可塑性，提供了对正常大脑发育的关键洞察， Rho GTP酶信号改变与认知/情绪障碍之间的联系，以及可能的治疗方法。

PROJECT SUMMARY Neural circuit formation and information processing in the brain require precise control of the development and remodeling of actin-rich dendritic spines and the excitatory synapses they house. Dynamic regulation of AMPA- and NMDA-type glutamate receptors, which mediate fast excitatory synaptic transmission and synaptic plasticity, respectively, is a key aspect of this control. Synaptic pathology characterizes many brain disorders including intellectual disabilities, autism, bipolar disorder, depression, and Alzheimer's disease. Thus, uncovering the mechanisms that control spine/synapse development and glutamate receptor regulation will provide critical insights into brain function and disease. Rho GTPases are master regulators of spine/synapse development and remodeling. Rac1 promotes spine/synapse formation, growth and maintenance, whereas RhoA suppresses these processes; both also play pivotal roles in synaptic plasticity. Proper function of Rho GTPases requires exquisite spatiotemporal control and disruption of this regulation results in numerous brain disorders. Rho GTPases are activated by guanine nucleotide exchange factors (GEFs) and inhibited by GTPase activating proteins (GAPs). However, remarkably little is known about how these GEFs/GAPs shape spatiotemporal Rac1/RhoA activation patterns and effector responses that direct the formation of neural circuits in brain. We identified the Rac1-GEF Tiam1 as a critical regulator of dendrite, spine, and synapse de- velopment, demonstrating that it couples synaptic receptors to Rac1 activation and actin cytoskeletal remodeling in cultured hippocampal neurons. In the last grant cycle, we made the surprising discovery that Tiam1 binds to the Rac1-GAP/RhoA-GEF Bcr and that this GEF/GAP complex is required to precisely regulate synaptic Rac1 signaling and excitatory synapse formation. Bcr is linked to bipolar disorder and learning and behavioral deficits, whereas altered Tiam1 expression is seen in patients with depression and Down syndrome. We hypothesize that Tiam1/Bcr cooperate to control the activation dynamics and signaling specificity of Rho GTPases, which is required in vivo for proper spine/synapse development, NMDAR trafficking/function, learning, and mood regulation. To test this, we propose to: (1) identify the roles of Tiam1 and closely related Tiam2 in shaping spine/synapse development in vivo and the specific pathways that mediate their effects; and (2) elucidate the mechanisms by which Tiam1/Bcr control NMDARs in synaptic plasticity, learning and mood regulation. We will use a multidisciplinary approach involving mouse genetics, time-lapse live-cell and in vivo two-photon imaging, Förster Resonance Energy Transfer (FRET), electrophysiology, biochemistry, molecular and cellular biology, and behavioral analyses. Our findings will elucidate key mechanisms that control Rho GTPase-dependent synaptic development/plasticity, providing critical insight into normal brain development, the connection between altered Rho GTPase signaling and cognitive/mood disorders, and potential treatments.