Fundamental Mechanisms of GPR56 Activation and Regulation

描述（由申请人提供）：大脑皮层发育异常可导致一系列不同的神经发育障碍，包括自闭症和自闭症综合征。对于复杂的多基因疾病（如自闭症）相关的皮质发育失调的新见解，可以从对简单的单基因疾病的研究中获得，这些疾病也植根于大脑皮质的异常发育。其中一种是双侧额顶叶多微脑回症（BFPP），患者表现出皮质连接紊乱，导致严重的认知异常。BFPP是一种常染色体隐性遗传综合征，由孤儿受体GPR 56突变引起，GPR 56是G蛋白偶联受体粘附家族的一员，但对其了解甚少。GPR 56具有极大的胞外氨基末端（NT），其与粘附蛋白具有同源性，并且已显示在受体加工期间被切割，但仍保持与受体的七跨膜（7 TM）区结合。然而，GPR 56信号传导激活和调节的基本机制目前尚不清楚。我们的初步研究表明，野生型GPR 56在异源细胞中的表达导致Rho和b-连环蛋白/Tcf信号传导的强烈激活，并且大GPR 56-NT的截短导致受体被有效地运输到质膜，但不再激活上述信号传导途径。我们假设GPR 56-NT与粘附伴侣的相互作用促进受体信号传导，并将通过阐明控制受体活性的GPR 56-NT的结构决定因素来探索这一想法。我们进一步假设，GPR 56信号是高度调节的细胞质激酶，抑制蛋白和PDZ支架，并将探讨这些可能性，使用磷酸化试验，成像研究，免疫共沉淀实验，siRNA敲低方法在转染细胞和培养的皮层神经元的组合。从这些研究中获得的知识将为GPR 56在人类疾病中的作用以及调节大脑皮层发育的基本机制提供新的见解。公共卫生相关性：大脑皮层发育异常会导致一系列不同的神经发育障碍，包括自闭症和自闭症综合征。为了阐明导致此类疾病的因素，我们建议研究孤儿受体GPR 56的基本特性，该受体被认为在皮质发育中起关键作用，因为GPR 56的突变会导致一种罕见的遗传疾病，其中大脑皮质的发育严重扭曲。从这些集中在GPR 56正常功能的研究中获得的知识将为GPR 56在人类疾病中的作用以及调节大脑皮层发育的基本机制提供新的见解。

DESCRIPTION (provided by applicant): Abnormalities in the development of the cerebral cortex can lead to a range of distinct neurodevelopmental disorders, including autism and Asperger's syndrome. Fresh insights into the dysregulation of cortical development associated with complex polygenetic disorders such as autism can be gained from studies on simpler monogenetic disorders that also are rooted in abnormal development of the cerebral cortex. One such disorder is bilateral frontoparietal polymicrogyria (BFPP), a condition in which patients exhibit disordered cortical connectivity resulting in profound cognitive abnormalities. BFPP is an autosomal recessive syndrome that results from mutations in the orphan receptor GPR56, which is a member of the poorly-understood adhesion family of G protein-coupled receptors. GPR56 possesses an extremely large extracellular amino-terminus (NT) that has homology to adhesion proteins and has been shown to be cleaved during receptor processing but yet remain associated with the seven-transmembrane (7TM) region of the receptor. However, the fundamental mechanisms of the activation and regulation of GPR56 signaling are presently unknown. Our preliminary studies have shown that expression of wild-type GPR56 in heterologous cells results in strong activation of Rho and b-catenin/Tcf signaling, and that truncation of the large GPR56-NT results in a receptor that is trafficked efficiently to the plasma membrane but no longer activates the aforementioned signaling pathways. We hypothesize that interaction of the GPR56-NT with an adhesion partner promotes receptor signaling, and will explore this idea by elucidating the structural determinants of the GPR56-NT that control receptor activity. We furthermore hypothesize that GPR56 signaling is highly regulated by cytoplasmic kinases, arrestins and PDZ scaffolds, and will explore these possibilities using a combination of phosphorylation assays, imaging studies, co-immunoprecipitation experiments, and siRNA knockdown approaches in both transfected cells and cultured cortical neurons. Knowledge gained from these studies will provide fresh insights into the role of GPR56 in human disease and also into the fundamental mechanisms that regulate the development of the cerebral cortex. PUBLIC HEALTH RELEVANCE: Abnormalities in the development of the cerebral cortex can lead to a range of distinct neurodevelopmental disorders, including autism and Asperger's syndrome. To shed light on the factors that contribute to such diseases, we propose to study the fundamental properties of the orphan receptor GPR56, which is believed to play a key role in cortical development since mutations in GPR56 cause a rare genetic disorder in which the development of the cerebral cortex is grossly distorted. Knowledge gained from these studies focused on the normal function of GPR56 will provide fresh insights into the role of GPR56 in human disease and also into the fundamental mechanisms that regulate the development of the cerebral cortex.