Regulation of MET Expression in Autism Disorder and Forebrain Ontogeny

描述(由申请人提供)： 这项拟议的研究有两个相关的长期目标。首先，我们的目标是确定人类MET启动子中与自闭症谱系障碍(ASD)脆弱性相关的功能变体的转录调节因子。其次，我们计划确定小鼠Met受体酪氨酸激酶影响前脑个体发育的机制。根据疾病控制中心的数据，ASD是一种严重的神经发育障碍，在美国的患病率为每150人中就有1人。最近，我们发现人类MET基因5‘转录调控区的单核苷酸多态(SNP)(rs1858830等位基因’C‘)与ASD密切相关(P=5x10-6)，并可减少转录因子(TF)结合和基因转录。人类遗传学研究在ASD和基础发育神经生物学方面的数据趋同表明，MET信号是前脑回路组装的重要组成部分，调节失调导致功能病理生理。我们将首先确定调节MET表达的转录因子。我们的初步筛选显示了29个假定的TF，我们将通过EMSA和芯片分析进行验证。过表达和siRNA将被用来在体外改变MET调节因子的水平，以描绘它们在调节MET转录中的单独和集体作用。接下来，我们将在小鼠中使用原位杂交来建立Met及其调控因子在脑发育过程中的时空表达。这些图谱对于探索在皮质发育过程中调节Met转录的功能相互作用至关重要。最后，我们将使用一种新的技术策略，在胚胎小鼠的宫内电穿孔，操纵LHX2，一个假定的蛋氨酸调节因子，在大脑发育过程中的表达。这些操作的双重目的是揭示体内TF干扰对Met表达的影响，并确定随后对皮质发育的影响。自闭症谱系障碍(ASD)是一种严重的神经发育障碍，影响大量个人及其家庭。我们已经确定了MET基因的一个变化，它改变了它被调控的方式，并使ASD的风险增加了一倍以上。已知MET会影响大脑发育，我们正在研究MET在大脑中正常调节的方式，以及调节失调可能如何导致大脑发育改变和ASD易感性增加。我们对MET调控的关注与许多健康问题高度相关，因为该基因的过度表达是高侵袭性恶性肿瘤的特征，我们的工作表明，表达不足可能是ASD的危险因素。

DESCRIPTION (provided by applicant): There are two related long-term goals of the proposed research. First, we aim to determine the transcriptional regulators of a functional variant in the human MET promoter that is associated with autism spectrum disorder (ASD) vulnerability. Second, we plan to define the mechanisms through which the mouse Met receptor tyrosine kinase influences forebrain ontogeny. ASD is a significant neurodevelopmental disorder, with a prevalence of 1 in 150 in the United States, according to the Centers for Disease Control. Recently, we discovered that a single nucleotide polymorphism (SNP) in the 5' transcriptional regulatory region of the human MET gene (rs1858830 allele 'C') is strongly associated with ASD (P=5x10-6), and also reduces transcription factor (TF) binding and gene transcription. The convergence of data from human genetics studies in ASD and basic developmental neurobiology suggests that MET signaling is an important component for the assembly of forebrain circuits, with dysregulation leading to functional pathophysiology. We will first identify TFs that regulate MET expression. Our preliminary screen revealed 29 putative TFs, which we will validate by EMSA and ChIP assays. Over-expression and siRNA will be used to alter levels of MET-regulating TFs in vitro to delineate their individual and collective roles in regulating MET transcription. Next, we will use in situ hybridization in the mouse to establish the spatial and temporal expression of Met and its regulators during brain development. These maps are crucial for probing functional interactions that regulate Met transcription during cortical development. Finally, we will use a novel technical strategy, in utero electroporation in embryonic mice, to manipulate the expression of Lhx2, a putative Met-regulating TF, during brain development. The dual purpose of these manipulations is to reveal the effects of TF perturbation on Met expression in vivo and to define the ensuing impact on cortical development. RELEVANCE Autism spectrum disorder (ASD) is a serious neurodevelopmental disorder that impacts a large number of individuals, and also their families. We have identified a change in the MET gene that alters the way in which it is regulated and more than doubles the risk of ASD. MET is known to influence brain development, and we are studying the ways in which it is normally regulated in the brain, and how dysregulation may lead to altered brain development and increased ASD susceptibility. Our focus on regulation of MET is highly relevant to many health issues, as over-expression of the gene is characteristic of highly invasive malignancies, and our work shows that under-expression may be a risk factor for ASD.