Axonal FMRP in Synaptic Development

项目摘要 这个项目将解决神经发育过程中突触发育异常的问题。 精神错乱。我们的总体假设是，错乱的突触粘连和延迟的功能组装 突触小泡损害突触前终末的形成和生理成熟，从而触发 随之而来的突触连接和功能发育缺陷。我们将在Fragile X中测试这一假设 综合征(FXS)，自闭症和智力残疾的一种主要可遗传形式，由 脆性X智力低下蛋白(FMRP)。实验观察将利用进化上保守的 在活体细胞自主特性的鸡胚中容易接近的末端。 我们将追求两个具体目标来检验从我们的初步研究中得出的几个重要假设。 ·在特定目标1中，我们将确定FMRP调节的突触黏附在突触前的作用 终端编队。我们假设轴突FMRP促进终末的形成、稳定和 通过发育轮廓突触粘连的选择性回缩。为了检验这一假设，我们将 使用特定细胞组和时间控制的基因操作与活体内活着相结合 FMRP介导的轴突运输与蛋白质翻译在动态中确切作用的成像研究 码头周转量。我们还将确定FMRP调节的突触黏附元件在发育过程中 并评估纠正这些元件对FMRP丢失诱导的突触前和 轴突改变。 ·在具体目标2中，我们将确定FMRP调节的突触素(SYT)在功能 突触前终末的成熟。Syt1/2是SVS上触发囊泡融合的主要钙感受器 和神经递质的释放。我们假设FMRP通过以下方式调节突触前功能成熟 控制新生终端中Syt2的及时上调。为了检验这一假设，我们将确定 Syt2基因表达对FMRP缺失所致突触前蛋白--SV活性缺失的影响 机械和谷氨酸释放。我们还将确定突触粘连之间的相互作用 使用救援研究，在FMRP控制下进行规范和SV组装。 总之，这些结果将确定突触表型缺陷的来源，这是神经发育的标志 精神错乱。这一知识至关重要，因为它将有助于建立一个敏感的时间窗口，并确定 预防或至少减少FXS和FXS中突触缺陷进展的新候选治疗方法 其他神经发育障碍。

Project Summary This project will address the question of how abnormal synaptic development emerges in neurodevelopmental disorders. Our overall hypothesis is that disorganized synaptic adhesion and delayed functional assembly of synaptic vesicles (SVs) impair the formation and physiological maturation of presynaptic terminals, which triggers subsequent developmental deficits in synaptic connectivity and function. We will test this hypothesis in Fragile X syndrome (FXS), a leading inheritable form of autism and intellectual disability caused by functional loss of Fragile X mental retardation protein (FMRP). Experimental observations will utilize the evolutionally conserved endbulb terminals that are readily accessible for in vivo cell-autonomous characterizations in chicken embryos. We will pursue two specific aims to test several important hypotheses derived from our preliminary studies. · In Specific Aim 1, we will determine the role of FMRP-regulated synaptic adhesion in presynaptic terminal formation. We hypothesize that axonal FMRP promotes terminal formation, stabilization, and selective retraction through developmentally profiled synaptic adhesion. To test this hypothesis, we will use cell-group specific and temporally-controlled genetic manipulations combined with in vivo live imaging to identify the exact actions of FMRP-mediated axon transport vs. protein translation in dynamic terminal turnover. We will also identify FMRP-regulated synaptic adhesion elements in developing terminals and assess the effects of correcting these elements on FMRP loss-induced presynaptic and axon alterations. · In Specific Aim 2, we will determine the role of FMRP-regulated synaptotagmin (Syt) in functional maturation of presynaptic terminals. Syt1/2 are primary calcium sensors on SVs that trigger vesicle fusion and neurotransmitter release. We hypothesize that FMRP regulates presynaptic functional maturation by controlling the timely upregulation of Syt2 in nascent terminals. To test this hypothesis, we will determine the effects of expressing Syt2 on FMRP loss-induced deficits in SV activity, presynaptic protein machinery, and glutamate release. We will also determine the interplay between synaptic adhesion regulation and SV assembly under FMRP control using rescue studies. Together, these results will identify an origin of defective synaptic phenotypes, a hallmark of neurodevelopmental disorders. This knowledge is of vital importance because it will help establish a sensitive time window and identify novel therapeutic candidates for preventing, or at least reducing, the progress of synaptic deficits in FXS and other neurodevelopmental disorders.