High-throughput Modeling of Autism Risk Genes using Zebrafish - DIVERSITY SUPPLEMENT

项目总结 自闭症谱系障碍(ASD)是由环境因素和遗传因素共同引起的，具有遗传性 贡献率估计为60%-80%。数十个增加ASD风险的基因已经被发现，其中大多数是基于 新生突变，但这些突变预计只占ASD病例的15%-20%。因此， 据预测，ASD的大部分遗传因素是由常见和罕见的遗传变异造成的，但 几乎没有这样的基因被发现。最近，利用全基因组测序，我们报道了全基因组 60个ASD风险基因的证据，其中26个是ASD的新基因，信号来自遗传和新生 蛋白质截断或错义突变。大多数这些基因的功能是未知的，所以一个关键的和 必要的下一步是使用模型来探索它们对神经发育和神经元功能的影响 有机体。目前将遗传风险因素转化为表型、机制和治疗的速度是 部分受限于体内哺乳动物模型系统的低效，这使得它们不适用于创建 并对大量突变品系进行行为测试。在这里，我们利用斑马鱼，它占据了一个利基 作为一种脊椎动物模型，具有适合活体筛选和机制理解的特征，包括 保守但较小的脊椎动物大脑，与自闭症相关的行为，以及相对于哺乳动物的成本效益 模特们。虽然斑马鱼不能概括ASD，并且在模拟人类疾病方面有局限性，但 新出现的文献支持这一概念，即这是一个有用的模型来研究基因的功能，有助于 ASD风险。与其一次只评估一个自闭症风险基因，我们将加速向机械化方向发展 通过高通量分析和分析了解。在家长的资助中，我们建议使用全脑 钙成像研究斑马鱼ASD危险基因突变体幼体阶段的神经元网络特性 发展。这个多样性补充应用程序描述了一个实验性和概念性的职业生涯 研究生的发展计划，其实验目标是(1)建立一个全脑系统 斑马鱼幼鱼在呈现虚拟社交线索时的钙成像，以及(2)使用该系统识别 斑马鱼ASD危险基因突变体与野生型对照的神经元网络特性比较 社交暗示。这项实验计划通过描述反应中的大脑状态，直接与父母的资助有关 在斑马鱼第一次表现出社会行为的幼鱼发育阶段的社会线索。这些 实验与父母拨款中描述的实验是分开的，但又是协同的。团结在一起， 父母补助和多样性补充有可能确定解释 在斑马鱼中观察到的包含ASD风险基因突变的行为表型。

PROJECT SUMMARY Autism spectrum disorder (ASD) is caused by both environmental and genetic factors, with the genetic contribution estimated at 60-80%. Dozens of genes that increase risk for ASD have been identified, most based on de novo mutations, but these mutations are predicted to account for only 15-20% of ASD cases. Thus, the majority of the genetic contribution to ASD is predicted to result from common and rare inherited variation, but few such genes have been identified. Recently, using whole genome sequencing, we reported genome wide evidence for >60 ASD risk genes, 26 of them novel for ASD, with signals derived from inherited and de novo protein truncating or missense mutations. The functions of most of these genes are unknown, so a crucial and necessary next step is to explore their impact on neurodevelopment and neuronal function using a model organism. The current pace of translating genetic risk factors into phenotypes, mechanisms and therapies is limited in part by inefficiencies with in vivo mammalian model systems, which makes them impractical for creating and behaviorally testing large numbers of mutant lines. Here, we leverage the zebrafish, which occupies a niche as a vertebrate model with features amenable to both in vivo screening and mechanistic understanding, including a conserved yet small vertebrate brain, behaviors relevant to ASD, and cost-effectiveness relative to mammalian models. While the zebrafish cannot recapitulate ASD and has limitations for modeling a human disorder, an emerging literature supports the notion that it is a useful model to study the functions of genes that contribute to ASD risk. Rather than assess ASD-risk genes one at a time, we will accelerate progress towards mechanistic understanding via high-throughput assays and analyses. In the parent grant, we proposed to use whole-brain calcium imaging to study neuronal network properties of zebrafish ASD risk gene mutants at the larval stage of development. This diversity supplement application describes an experimental and conceptual career development plan for a graduate student whose experimental goals are to (1) establish a system for brain-wide calcium imaging of juvenile zebrafish during presentation of virtual social cues, and (2) use this system to identify neuronal network properties of zebrafish ASD risk gene mutants compared to wild-type controls in response to social cues. This experimental plan directly relates to the parent grant by characterizing brain states in response to social cues at the juvenile stage of development, when zebrafish first show social behaviors. These experiments are separate from, yet synergize with, the experiments described in the parent grant. Together, the parent grant and diversity supplement have the potential to identify neuronal mechanisms that explain the behavioral phenotypes observed in zebrafish that contain mutations in ASD risk genes.