Characterizing patient-specific TBR1 mutations: Understanding a master regulator of autism risk.

项目摘要/摘要 自闭症谱系障碍(Asd)是一种常见的神经发育障碍，其特征是 社会交流和受限、重复的行为。到目前为止，有效的治疗方法有限 帮助个人管理ASD的核心症状。ASD风险背后的遗传因素的多样性 强调需要将我们的重点从一刀切的所有疗法转移到更定制、更个性化的疗法上。 最近，下一代测序(NGS)使研究人员能够识别从头开始(新出现在 儿童)许多重要的大脑发育基因的功能突变，包括与 无血缘关系的儿童中存在多种突变。这些发现为反复突变的基因提供了强有力的证据。 在ASD风险中起着重要作用，并表明单个基因的杂合性破坏对于 大脑发育足以导致自闭症。转录因子tbr1，它是一种“主调控因子”。 在大脑发育中，是一种特别令人感兴趣的基因。在大约0.2%的ASD儿童中，TBR1发生了突变， 使其成为最常见的风险因素之一。利用生物网络方法进行计算分析 这表明，尽管遗传复杂，但在特定的发育窗口和 大脑区域可能在ASD的基因亚群中发挥作用。具体地说，高信心风险的共同表达 基因在皮质发育的胎儿中期阶段汇聚，在这一阶段，tbr1被认为在 深层谷氨酸能皮质投射神经元的分化、迁移和功能。此外，它是 现在清楚的是，TBR1还与其他三分之一的高信度自闭症风险基因结合并调节，使其成为一种 至少一种新出现的常见自闭症病因学的潜在“大师”。 评估特定突变的功能后果是验证和 了解基因和表型之间的因果关系并设计合理的目标 治疗/干预。我们假设，单个TBR1拷贝的丢失会扰乱人类大脑皮层 通过改变TBR1调节的网络进行发育，这是正确的神经元识别和迁移所必需的。 此外，在ASD风险的关键发育窗口期间干扰TBR1或其目标基因定义了 通向自闭症的常见途径。我们先前证实从头基因突变严重影响定位和 突变的tbr1蛋白对靶基因的调控能力。在这里，我们将解决我们目前在 利用尖端基因组了解患者特有的TBR1突变如何影响发育中的神经元 编辑、功能基因组学和利用患者来源的诱导多能性的补充模型 干细胞(IPSCs)转化为类前脑器官(Aim 1)和小鼠遗传学(Aim 2)。这些研究 将提供一个史无前例的视角来研究TBR1患者特异性突变对神经元的影响 在大脑皮层发育期间。此外，这种IPSC/小鼠遗传平台还可以扩展到其他风险 并成为设计合理的针对自闭症及相关疾病的靶向治疗/干预措施的基础。

PROJECT SUMMARY/ABSTRACT Autism spectrum disorder (ASD) is a common neurodevelopmental disorder characterized by impaired social communication and restricted, repetitive behaviors. As of now, there are limited effective therapies to help individuals manage the core symptoms of ASD. The diversity of genetic factors underlying ASD risk highlights the need to shift our focus from one-size fits all therapeutics to more tailored, individualized therapies. Recently, next-generation sequencing (NGS) has enabled researchers to identify de novo (newly appearing in the child) loss of function mutations in many important brain development genes, including genes with in multiple mutations in unrelated children. These findings provide strong evidence for recurrently mutated genes playing a significant role in ASD risk and indicate that heterozygous disruption of a single gene essential for brain development is sufficient to cause autism. TBR1, a transcription factor that serves as a `master regulator' in brain development, is one such gene of particular interest. TBR1 is mutated in ~0.2% of children with ASD, making it one of the most common risk factors. Computational analyses using biologic network approaches suggest that, despite the genetic complexity, converging biology at particular developmental windows and brain regions may be at play in genetic subsets of ASD. Specifically, the co-expression of high confidence risk genes converge at midfetal stages of cortical development, where TBR1 is thought to play a key role in the differentiation, migration, and function of deep layer glutamatergic cortical projection neurons. Moreover, it is now clear that TBR1 also binds to and regulates ~1/3 of other high confidence autism risk genes, making it a potential `master regulator' of at least one emerging common autism etiology. Evaluating the functional consequences of specific mutations represents a critical step in validating and understanding the causal link between genotype and phenotype and designing rational targeted therapies/interventions. We hypothesize that loss of a single copy of TBR1 disrupts human cortical development by altering the TBR1-regulated network required for proper neuronal identity and migration. Moreover, disrupting TBR1 or its target genes during this critical developmental window of ASD risk define a common route to ASD. We previously confirmed that de novo mutations severely impacted the localization and ability of mutant TBR1 proteins to regulate target genes. Here, we will address the current gaps in our knowledge of how patient-specific TBR1 mutations affect developing neurons by utilizing cutting-edge genome editing, functional genomics, and complementary models that leverage patient-derived induced pluripotent stem cells (iPSCs) converted to forebrain-like organoids (Aim 1) and mouse genetics (Aim 2). These studies will provide an unprecedented view into the consequences of TBR1 patient-specific mutations on neurons during cortical development. Moreover, this iPSC/mouse genetics platform can be expanded to other risk genes and be the basis for designing rational targeted therapies/interventions for ASD and related disorders.