Defining links between an intellectual disability associated RNA-binding protein and planar cell polarity in neurodevelopment

项目概要： 智力残疾影响着世界人口的1-3%;这些人在适应能力方面存在缺陷。 功能需要持续的支持，以执行活动，如穿衣，阅读，并解释 别人的意图。这种支持的成本，为一个单一的美国病人，大约是1 -2百万美元，整个 他或她的寿命1.智力残疾的病理生理学和病因学一直难以阐明， 偶然突变的异质性2.尽管有这种复杂性，与智力残疾有关的突变往往 在与神经系统发育、细胞代谢和微管相关的途径中积累 运动和轴突运输3.此外，单基因形式的智力残疾提供了直接的洞察力， 导致智力残疾的缺陷细胞生物学过程。我们的实验室发现， 普遍表达的锌指，聚腺苷RNA结合蛋白，ZC 3 H14，与一种形式的 单基因、非综合征型常染色体隐性智力残疾4.我们发明了一种果蝇 黑腹果蝇模型，以研究dNab 2的作用，dNab 2是人ZC 3 H14的果蝇直系同源物。dNab 2结果丢失 在神经元、存活和运动表型方面。重要的是，许多表型可以通过 人ZC 3 H14的转基因表达仅在神经元中表达，这意味着果蝇的功能保守 以及使用dNab 2模拟ZC 3 H14功能的可行性。我们的苍蝇模型生成的数据表明 dNab 2缺失在神经发育中至关重要，并且它可能调节神经元中的基因表达。 然而，dNab 2在神经元中的mRNA靶点的身份及其调节这些靶点的机制尚不清楚。 目标是知识方面的主要差距。初步数据表明，dNab 2在功能上与多种 平面细胞极性（PCP）途径的组成部分。PCP是Wnt信号传导的非规范分支， 调节神经系统中的轴突导向和体细胞组织中的组织极化5-8。所以我会 直接检验dNab 2调节PCP组分以控制神经突延伸和引导的假设 在神经发育过程中。该项目的具体目标是：1）确定基因dNab 2：五氯苯酚的相互作用， 两种神经发育背景，2）利用系统水平的蛋白质组学方法来鉴定dNab 2调节的 研究dNab 2和PCP通路之间的物理和功能相互作用 体内RNA。目的1利用果蝇遗传工具评估两种果蝇中与dNab 2的功能性遗传相互作用 神经元细胞类型。目的2使用网络分析，一个独特的果蝇蛋白质组数据集，以确定dNab 2 调控途径。目的3利用技术来评估RNA定位和与dNab 2的物理相互作用。 这些目标的成功完成将提供深入了解dNab 2如何调节局部基因表达， 影响神经发育，从而支持我们广泛的，长期的目标，定义细胞生物学和 ZC 3 H14/dNab 2相关神经发育缺陷的分子机制

PROJECT SUMMARY: Intellectual disability affects 1–3% of the worldwide population; these individuals have deficits in adaptive functioning necessitating ongoing support to perform activities such as dressing, reading, and interpreting the intentions of others. The cost of this support, for a single US patient, is approximately $1–2 million throughout his or her lifespan1. The pathophysiology and etiology of intellectual disability has been difficult to elucidate due to the heterogeneity in casual mutations2. Despite this complexity, mutations linked to intellectual disability tend to accumulate in pathways relating to nervous system development, cellular metabolism, and microtubule based movement and axonal transport3. Additionally, monogenic forms of intellectual disability provide direct insight into defective cell biological processes that underlie intellectual disability. Our lab found that mutations in a ubiquitously expressed zinc-finger, polyadenosine RNA-binding protein, ZC3H14, are linked to a form of monogenic, non-syndromic autosomal recessive intellectual disability4. We developed a Drosophila melanogaster model to investigate the role of dNab2, the fly ortholog to human ZC3H14. Loss of dNab2 results in neuronal, survival, and locomotive phenotypes. Importantly, many of the phenotypes can be rescued by transgenic expression of human ZC3H14 exclusively in neurons, implying a conservation of function from flies to humans and feasibility in using dNab2 to model ZC3H14 function. Data generated from our fly model suggests that dNab2 loss is critical in neurodevelopment, and that it may be regulating gene expression in neurons. However, the identity of mRNA targets of dNab2 in neurons and the mechanism by which it regulates these targets are key gaps in knowledge. Preliminary data suggest that dNab2 interacts functionally with multiple components of the planar cell polarity (PCP) pathway. PCP is a non-canonical branch of Wnt signaling that regulates axon guidance in the nervous system and tissue polarization in somatic tissue 5–8. Therefore, I will directly test the hypothesis that dNab2 regulates PCP components to control neurite extension and guidance during neurodevelopment. The Specific Aims of this project are: 1) define genetic dNab2:PCP interactions in two neurodevelopmental contexts, 2) utilize a systems-level proteomic approach to identify dNab2 regulated pathways in the brain, and 3) examine physical and functional interactions between dNab2 and PCP pathway RNAs in vivo. Aim 1 utilizes Drosophila genetic tools to assess functional genetic interactions with dNab2 in two neuronal cell types. Aim 2 uses network analyses, of a unique Drosophila proteomic dataset, to identify dNab2 regulated pathways. Aim 3 utilizes techniques to assess RNA localization and physical interaction with dNab2. Successful completion of these aims will provide insight into how dNab2 regulates local gene expression to impact neurodevelopment, and thus support our broad, long-term objective of defining the cell biological and molecular mechanisms underlying ZC3H14/dNab2 related neurodevelopmental defects.