Cellular and molecular mechanisms disrupted in 22q13 deletion syndrome and autism

编码突触蛋白的基因突变和脑功能连接受损正在出现 作为与自闭症谱系障碍(ASD)相关的常见缺陷。然而，突触是如何突变的 蛋白质在细胞和分子水平上影响人类神经元的特性，从而导致大脑异常 联系仍然是一个重要的悬而未决的问题。解决这个问题对于 了解ASD的病因和病理，开发新的有效的治疗策略 对病人来说。PI先前证明了SHANK3缺陷的人类皮质神经元来源于 自闭症患者22q13缺失综合征患者产生的诱导多能干细胞(IPSCs) 严重损害内在兴奋性和兴奋性突触传递。然而，这两个人怎么 表型会发展并影响大脑中神经元的连通性，目前尚不清楚。这样做的主要目标是 该项目旨在阐明负责突触和神经系统发育的细胞和分子机制。 SHANK3基因缺陷的人类神经元的连接缺陷。SHANK3是一种支架蛋白，表达于 自闭症患者和自闭症患者中经常发现的兴奋性突触突变或缺失 智力残疾。该项目的中心假设是ShanK3缺陷人类的突触缺陷 神经元的发育是由于电活动的增加和活动介导的减弱和消除 兴奋性突触。这一假设得到了PI获得的初步数据的有力支持 实验室。为了检验这一假设，制定了以下具体目标：1)确定提升的角色 SHANK3缺陷型人类神经元突触缺陷形成过程中的电活动；2)确定 ARC在SHANK3缺陷型人类神经元突触缺陷形成中的作用；以及3)决定如何 在活体中，人类神经元中SHANK3的缺失会影响这些神经元的突触输入。在这些目标下， 从新的、精确的基因工程干细胞系产生的人类皮质神经元的特性将 在体外和植入后使用电生理学、成像和生化技术进行研究 进入老鼠的大脑。这项拟议的研究意义重大，因为预计它将大大推进 了解自闭症和智力障碍的分子、细胞和电路机制 为残疾患者制定新的治疗策略并指导其发展。

Mutations in genes encoding synaptic proteins and impaired functional brain connectivity are emerging as common deficits associated with autism spectrum disorders (ASDs). However, how mutated synaptic proteins affect the properties of human neurons at the cellular and molecular levels to cause abnormal brain connections remains an important unanswered question. Addressing this question is essential for understanding the etiology and pathology of ASDs and developing novel and effective therapeutic strategies for patients. The PI previously demonstrated that SHANK3-deficient human cortical neurons derived from induced pluripotent stem cells (iPSCs), generated from 22q13 deletion syndrome patients with autism, have severely impaired intrinsic excitability and excitatory synaptic transmission. However, how these two phenotypes develop and affect neuronal connectivity in the brain remain unknown. The main goal of this project is to elucidate the cellular and molecular mechanisms responsible for development of synaptic and connectivity deficits in SHANK3-deficient human neurons. SHANK3 is a scaffolding protein expressed at excitatory synapses that have been frequently found to be mutated or deleted in individuals with autism and intellectual disability. The central hypothesis of this project is that synaptic deficits in SHANK3-deficient human neurons develop as a result of elevated electrical activity and activity-mediated weakening and elimination of excitatory synapses. This hypothesis is strongly supported by the preliminary data obtained in the PI's laboratory. The following Specific Aims are formulated to test this hypothesis: 1) Determine the role of elevated electrical activity in development of synaptic deficits in SHANK3-deficient human neurons; 2) Determine the role of ARC in development of synaptic deficits in SHANK3-deficient human neurons; and 3) Determine how loss of SHANK3 in human neurons impacts synaptic inputs onto these neurons in vivo. Under these aims, the properties of human cortical neurons generated from novel, precisely genetically-engineered stem cell lines will be investigated using electrophysiology, imaging, and biochemistry techniques in vitro and upon engraftment into the mouse brain. The proposed research is significant because it is expected to substantially advance understanding of the molecular, cellular, and circuitry mechanisms disrupted in autism and intellectual disability, and guide the development of novel therapeutic strategies for patients.