Local protein synthesis in presynaptic plasticity

项目总结 突触根据活动改变其功能和结构属性的能力，称为 突触可塑性，被认为是学习和记忆的分子基础，并严重依赖于 蛋白质合成。蛋白质合成的调节对于正常的突触功能是必不可少的，并在 突触病态疾病，如脆性X综合征(FXS)和自闭症谱系障碍(ASDS)。尽管多年来 研究表明，这些疾病仍然没有有效的治疗方法，给社会带来了巨大的负担，因为许多 患有FXS/ASD的成年人需要照顾者，无法找到工作。突触可塑性可能是由于 突触后受体改变或突触前神经递质释放的变化。虽然大多数研究都是 FXS专注于突触后可塑性机制，包括局部蛋白质合成，相对较少 了解FXS背景下长期突触前变化的机制。这个 脆性X智力低下蛋白(FMRP)在FXS中发生突变，是一种调节 依赖活性的局部蛋白质合成。FMRP在突触前表达，但它在长期 突触前可塑性从未被研究过。利用尖端成像技术，如超级 分辨率显微镜和双光子激光扫描显微镜(2PLM)与电生理相结合，这 提案将研究突触前蛋白的合成，以及FMRP对其活性依赖的调节 可塑性。将使用荧光非规范氨基酸标记(FUNCAT)来测量局部突触前 突触前可塑性背景下的蛋白质合成。蛋白质合成的突触前操纵将 能够研究导致神经递质释放的持久变化的特定细胞机制 在突触前。为了测试突触前FMRP的作用，将使用FMRP的条件敲除模型 专门删除突触前细胞中的蛋白质。双光子显微镜和电生理学将 实现对支配活动依赖的突触前结构和 功能可塑性。这一提议将揭示迄今为止未知的长期变化机制 神经递质的释放，这是一个以前被忽视的研究领域，可能会对 FXS等疾病的突触病理学。

PROJECT SUMMARY The ability of synapses to change their functional and structural properties in response to activity, called synaptic plasticity, is believed to act as the molecular basis of learning and memory and relies critically on protein synthesis. The regulation of protein synthesis is essential for normal synaptic function and is altered in synaptopathic diseases like Fragile X Syndrome (FXS) and Autism Spectrum Disorders (ASDs). Despite years of research, these diseases still have no effective treatment and place a large burden on society, since many adults with FXS/ASDs require a caregiver and cannot hold a job. Synaptic plasticity can occur as a result of postsynaptic receptor modifications or presynaptic changes in neurotransmitter release. While most studies of FXS have focused on postsynaptic plasticity mechanisms, including local protein synthesis, relatively little is known about the mechanisms underlying long-term presynaptic changes in the context of FXS. The Fragile X mental retardation protein (FMRP), mutated in FXS, is an mRNA binding protein that regulates activity-dependent local protein synthesis. FMRP is expressed presynaptically, but its role in long-term presynaptic plasticity has never been investigated. Utilizing cutting-edge imaging techniques like super- resolution microscopy, and two-photon laser scanning microscopy (2PLM) coupled with electrophysiology, this proposal will investigate presynaptic protein synthesis, and its regulation by FMRP, in activity-dependent plasticity. Fluorescent Noncanonical Amino Acid Tagging (FUNCAT) will be used to measure local presynaptic protein synthesis in the context of presynaptic plasticity. Presynaptic manipulations of protein synthesis will enable investigation of the cell-specific mechanisms that lead to enduring changes in neurotransmitter release at the presynapse. To test the role of presynaptic FMRP, a conditional knock out model of FMRP will be used to specifically delete the protein in the presynaptic cell. Two-photon microscopy and electrophysiology will enable real-time assessment of the mechanisms that govern activity-dependent presynaptic structural and functional plasticity. This proposal will uncover heretofore unknown mechanisms of long-term changes in neurotransmitter release, a previously neglected line of research, and likely generate new insights into the synaptic pathology of diseases like FXS.