The roles of Huntingtin Associated Protein 40 in Huntingtin functions and Huntingtons disease pathogenesis

亨廷顿相关蛋白 40 在亨廷顿功能和亨廷顿病发病机制中的作用

• 批准号: 10405375
• 负责人: Sheng Zhang
• 金额: $ 5.06万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10405375
• 关键词: Affect; Animal Model; Animals; Binding; Biological Assay; Biological Models; Brain; Cell physiology; Complex; Disease model; Drosophila genus; Drug Targeting; Etiology; Evaluation; Evolution; Foundations; Genetic; Genetic Models; Glutamine; Homologous Gene; Human; Huntington Disease; Huntington gene; Huntington protein; Lead; Mammalian Cell; Mammals; Molecular; Molecular Conformation; Mutate; Neurons; Pathogenesis; Patients; Phenotype; Physiological; Proteins; Proteomics; Regulation; Reporting; Role; Sampling; Testing; Toxic effect; fly; genetic testing; in vivo; multidisciplinary; mutant; novel therapeutics; polyglutamine

Project Summary/Abstract Huntington's disease (HD) is caused by an abnormal expansion of the glutamine tract (polyQ) in Huntingtin (HTT). A clear understanding on how endogenous HTT is regulated in vivo is critical both for elucidating HD etiology and for identifying effective drug targets. HTT has numerous reported HTT associated partners (HAPs) and is functionally implicated in a growing list of cellular processes. However, little is known how HTT itself is regulated and whether such regulation is altered in HD. We previously characterized the HTT homolog (dHtt) in model organism Drosophila. Given the significant functional conservation of HTT from the fly to mammals, we hypothesized that the core regulators of HTT likely are among the numerous known HAPs and should also be conserved in Drosophila. In a proteomic study for such conserved central regulators of HTT in Drosophila, we isolated dHap40, the fly homolog of HAP40, as the strongest dHtt interactor. Importantly, converging evidence from studies in multiple species all support that in vivo HTT protein normally exists in a complex with HAP40, and HAP40 binding stabilizes the conformation of HTT. Further, in samples from HD patients, a ~10- fold increase of the levels of endogenous HAP40 were observed as compared to controls. However, despite these findings, by now there is no reported functional study of HAP40 in any physiological settings, and its effect on HTT's normal functions and mutant HTT toxicity remains unclear. Our preliminary studies support the significantly conserved physical and functional interactions between HTT and HAP40, implying a highly important regulatory relationship that constrains their co-evolution from flies to humans. Our findings not only establish Drosophila as a relevant genetic model to study the physiological roles of HAP40, but also lead to our hypothesis that HAP40 is a conserved central regulator of HTT and potentially a critical modulator of mutant HTT toxicity. Using established assays and HD models in Drosophila and cultured mammalian cells, we will systematically test this hypothesis. In Aim 1, we will carry out a comprehensive phenotypic analyses of dhap40 gene and test its genetic interactions with dhtt, so as to obtain a first systematic evaluation of HAP40 in a physiological setting and clarify its relationship with HTT at whole-animal level. In Aim 2, we will systematically test whether HAP40 is a central regulator of HTT's subcellular dynamics and its diverse cellular functions, so as to elucidate its relationship with HTT at molecular and cellular levels. In Aim 3, taking advantage of the well- established HD models in Drosophila and mammalian neurons, we will rigorously interrogate the role of HAP40 on mutant HTT toxicity. From these multidisciplinary studies, we will obtain a first comprehensive evaluation on the physiological functions of HAP40, its effect on endogenous HTT functions and on HD pathogenesis. The results potentially lay foundation on novel therapeutic avenues against HD via HAP40.

项目概要/摘要 亨廷顿病 (HD) 是由亨廷顿蛋白中谷氨酰胺束 (polyQ) 的异常扩张引起的 （HTT）。清楚地了解内源性 HTT 在体内如何调节对于阐明 HD 至关重要 病因学和确定有效的药物靶点。 HTT 拥有众多报告的 HTT 关联合作伙伴 (HAP) 并且在功能上与越来越多的细胞过程有关。然而，人们对 HTT 本身的了解却很少。 监管以及这种监管是否在 HD 中发生改变。我们之前表征了 HTT 同系物 (dHtt) 在模式生物果蝇中。鉴于从果蝇到哺乳动物，HTT 具有显着的功能保守性， 我们假设 HTT 的核心调节器可能是众多已知的 HAP 之一，并且也应该 保存在果蝇中。在一项针对果蝇 HTT 保守中央调节因子的蛋白质组学研究中， 我们分离出 dHap40，即 HAP40 的果蝇同源物，作为最强的 dHtt 相互作用因子。重要的是，收敛 来自多个物种研究的证据都支持体内 HTT 蛋白通常存在于与 HAP40 和 HAP40 结合可稳定 HTT 的构象。此外，在 HD 患者的样本中，~10- 与对照相比，观察到内源性 HAP40 水平成倍增加。然而，尽管 这些发现，到目前为止还没有报道 HAP40 在任何生理环境下的功能研究，及其 对 HTT 正常功能和突变 HTT 毒性的影响仍不清楚。我们的初步研究支持 HTT 和 HAP40 之间的物理和功能相互作用显着保守，这意味着高度 重要的调控关系限制了它们从果蝇到人类的共同进化。我们的发现不仅 建立果蝇作为相关遗传模型来研究HAP40的生理作用，也导致我们 假设 HAP40 是 HTT 的保守中央调节剂，并且可能是突变体的关键调节剂 HTT 毒性。使用果蝇和培养的哺乳动物细胞中已建立的测定和 HD 模型，我们将 系统地检验这一假设。在目标1中，我们将对dhap40进行全面的表型分析 基因并测试其与 dhtt 的遗传相互作用，从而获得 HAP40 在 生理环境，并在整个动物水平上阐明其与 HTT 的关系。在目标 2 中，我们将系统地 测试 HAP40 是否是 HTT 亚细胞动力学及其多种细胞功能的中央调节器，因此 阐明其与 HTT 在分子和细胞水平上的关系。在目标 3 中，充分利用 在果蝇和哺乳动物神经元中建立HD模型后，我们将严格探究HAP40的作用 突变型 HTT 毒性。从这些多学科研究中，我们将获得对 HAP40 的生理功能、其对内源性 HTT 功能和 HD 发病机制的影响。这 结果可能为通过 HAP40 对抗 HD 的新治疗途径奠定基础。