Molecular mechanisms and genetic drivers of reciprocal genomic disorders

摘要 相互性基因组疾病（RGDs）涉及相同基因组的重复性微缺失和微重复， 片段RGDs由非等位基因同源重组（NAHR）介导，并且共同地在 神经发育障碍（NDD）和先天性异常最常见的复发性遗传原因 在人类身上。鉴于RGDs的影响通常在发育早期，这些疾病不成比例地 影响儿童，往往导致终身残疾。基因的发现， 因此，RGDs和构成这些疾病成分的基因将代表异常高的 一个优先的机制研究和治疗靶向跨越孟德尔和复杂的频谱目标 紊乱我们的初步数据表明，一个集成的体外和体内分子和计算 使用细胞和动物模型的基因组学方法可以识别与以下相关的分子特征： RGDs和异常表型和失调网络的遗传驱动因素。在这些研究中，我们将 首先确定与微缺失相关的基因表达谱和细胞表型， 在来自等基因诱导的神经衍生物中8-12个最普遍的RGD区域的微复制 多能干细胞（iPSC）系。我们将使用CRISPR/Cas9基因组编辑方法来实现这一目标， 最近开发的靶向侧翼节段复制和模仿NAHR介导的机制 人类（目标1）。然后，我们将寻找与RGD相关表型相关的特定基因， 在斑马鱼中进行高通量驱动基因筛选（目标2），以评估所有单个基因，并成对 RGD区域内的相互作用。在目标3中，我们将寻求验证这些预测的驱动因素，并确定 它们在不同神经元谱系和小鼠组织中的影响。因此，这些研究将遵循一个框架 我们的研究小组以前曾用于确定非复发性微缺失综合征的遗传驱动因素， 和若干区域指导方针区域，包括16p11.2区域指导方针，并采用创新办法和技术， 使我们能够大规模地进行这些研究，并比较不同RGDs的特征。最后，tehse 分析将确定人类中最常见的RGDs的分子特征，即驱动RGDs的基因。 这些签名的特定成分，它们的组织特异性，以及拯救最强的 通过体外和体内的剂量操作来识别特征。

ABSTRACT Reciprocal genomic disorders (RGDs) involve recurrent microdeletion and microduplicaton of identical genomic segments. RGDs are mediated by non-allelic homologous recombination (NAHR) and are collectively among the most common recurrent genetic causes of neurodevelopmental disorders (NDD) and congenital anomalies in humans. Given that the impact of RGDs is usually early in development, these disorders disproportionately affect children and often result in lifelong disabilities. Discovery of the genes the molecular consequences of RGDs and the genes that underlie components of these disorders would therefore represent exceptionally high priority targets for mechanistic studies and therapeutic targeting across a spectrum of Mendelian and complex disorders. Our Preliminary Data suggest that an integrated in vitro and in vivo molecular and computational genomics approach using cellular and animal modeling can identify molecular signatures associated with RGDs and the genetic drivers of aberrant phenotypes and dysregulated networks. In these studies, we will first define the gene expression profiles and cellular phenotypes associated with microdeletion and microduplication of the 8-12 most prevalent RGD regions in neural derivatives from isogenic induced pluripotent stem cell (iPSC) lines. We will accomplish this using a CRISPR/Cas9 genome editing approach we recently developed that targets the flanking segmental duplications and mimics NAHR-mediated mechanisms in humans (Aim 1). We will then seek the specific genes associated with RGD-associated phenotypes using high-throughput driver gene screening in zebrafish (Aim 2) to evaluate all individual genes and pairwise interactions within RGD regions. In Aim 3 we will then seek to validate these predicted drivers and determine their impact in diverse neuronal lineages and across mouse tissues. These studies will thus follow a framework our investigative team has previously used to identify genetic drivers of non-recurrent microdeletion syndromes and several RGD regions, including 16p11.2 RGD, and apply innovative approaches and technologies to enable us to conduct these studies at scale and compare signatures across RGDs. At their conclusion, tehse analyses will define the molecular signatures of the most common RGDs in humans, the genes that drive specific components of these signatures, their tissue specificity, and the capacity to rescue the strongest signatures through dosage manipulation in vitro and in vivo.