Protein Interaction Network Analysis to Test the Synaptic Hypothesis of Autism

项目总结/摘要 “使用蛋白质-蛋白质相互作用网络分析评估自闭症的突触假说” 细胞的蛋白质-蛋白质相互作用（PPI）网络被认为代表具有涌现网络特性的系统， 将各种输入的信号整合成协调的响应。对蛋白质复合物的更深入了解 突触将增强我们对正常大脑功能的了解， 包括自闭症申请人最近开发了一种新的技术，多重免疫沉淀， 流式细胞术（mIP-FCM），允许快速，同时，定量测量数百种蛋白质-蛋白质 从少量的生物样品中检测相互作用（PPI）。该提案涉及将mIP-FCM技术 以与自闭症相关的突触蛋白为目标，并研究蛋白质复合物的定量变化， 在生理和疾病相关条件下发生。蛋白质动态活性的观察 小鼠和人类谷氨酸突触的复合物将有助于更好地理解正常脑功能， 并可能突出自闭症相关基因的病理分子机制。 申请人提出了三个具体目标：1）产生靶向突触蛋白的多重IP-FCM测定， 将在K99部分的第一年完成; 2）在携带自闭症相关基因的小鼠中绘制突触PPI。 突变。这一部分将在K99/R 00过渡期间的第2-5年完成。3)人类突触PPI图谱 iPS神经元来源于自闭症和典型发育个体，在R 00阶段的3-5年完成 获奖名单这项提案的总体目标是确定在PPI网络的具体变化， 突触与不同的自闭症危险因素，并调查在多大程度上有自闭症的收敛 风险基因进入突触处的特定共同分子通路。 在K99阶段，申请人将：1）开发针对 神经元突触，并学习适合大型数据集的统计和可视化方法; 2）学习 管理研究团队，包括管理技术支持人员和学生，监督预算，整合 数据来自多个项目，并担任初级科学家的导师。申请人召集了一组 经验丰富的共同导师，以帮助这种职业发展培训。总体而言，这一培训将促进 实现申请人的长期目标，发展国际知名的独立研究 该项目专注于自闭症风险因素的分子机制。

Project Summary/Abstract "Evaluating the synaptic hypothesis of autism using protein-protein interaction network analysis" Protein-protein interaction (PPI) networks of a cell are thought to represent a system with emergent network properties, integrating signals for a variety of inputs into coordinated responses. A deeper understanding of protein complexes at the synapse will enhance our knowledge of normal brain function and may highlight new therapeutic targets for diseases, including autism. The applicant has recently developed a novel technology, multiplex immunoprecipitation measured by flow cytometry (mIP-FCM) that allows the rapid, simultaneous, quantitative measurement of hundreds of protein-protein interactions (PPIs) from small amounts of biological samples. This proposal involves translating mIP-FCM technology to target synaptic proteins relevant to autism, and investigating the quantitative changes in protein complexes that occur under physiological and disease-associated conditions. The observation of the dynamic activity of protein complexes at the mouse and human glutamate synapse will contribute to a greater understanding of normal brain function, and may highlight pathological molecular mechanisms of autism-associated genes. The applicant proposes three specific aims: 1) Generate a multiplex IP-FCM assay targeted to synaptic proteins, to be completed during year 1 of the K99 portion of the award; 2) Map synaptic PPIs in mice carrying autism-associated mutations. This portion will be completed in years 2-5, during the K99/R00 transition. 3) Map synaptic PPIs in human iPS neurons derived from autistic and typically developing individuals, to be completed during years 3-5 of the R00 phase of the award. The overall goal of this proposal is to define specific changes in the PPI network of the glutamatergic synapse associated with different autism risk factors, and to investigate the extent to which there is convergence of autism risk genes into specific common molecular pathways at the synapse. During the K99 phase of the award, the applicant will 1) Develop the mIP-FCM assay targeted to the glutamatergic synapse, and learn appropriate statistical and visualization methods for the large datasets; 2) Learn to manage a research team, including management of technical support staff and students, overseeing budgets, integrating data from multiple individual projects, and acting as a mentor of junior scientists. The applicant has assembled a group of experienced co-mentors to help with this career development training. Overall, this training will facilitate the achievement of the applicant's long-term goal to develop an internationally-renowned independent research program focused on the molecular mechanisms of autism risk factors.