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

亨廷顿病(HD)是由亨廷顿谷氨酰胺束(PolyQ)异常扩张引起的 (HTT)。清楚地了解内源性HTT在体内是如何调节的，对于阐明HD和HD都至关重要 用于病因学和确定有效的药物靶点。据报道，HTT拥有众多HTT相关合作伙伴(HAP) 并在功能上与越来越多的细胞过程有关。然而，人们对HTT本身知之甚少 以及房屋署是否改变了这方面的规定。我们以前描述了HTT同源基因(DHtt) 在模式生物果蝇中。鉴于HTT从苍蝇到哺乳动物的显著功能保守， 我们假设HTT的核心调节者很可能是众多已知的HAP之一，并且应该也 在果蝇身上是保守的。在对果蝇HTT这种保守的中枢调节因子的蛋白质组学研究中， 我们分离出HAP40的果蝇同源基因dHap40，作为最强的dHtt相互作用因子。重要的是，融合 来自多个物种的研究证据都支持体内HTT蛋白通常存在于与 HAP40和HAP40结合稳定了HTT的构象。此外，在HD患者的样本中，a~10- 与对照组相比，内源性HAP40水平增加了一倍。然而，尽管 这些发现，到目前为止，还没有关于HAP40在任何生理环境中的功能研究的报道，而且它的 对HTT的正常功能和突变的HTT毒性的影响尚不清楚。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著保守，这意味着 重要的调控关系，限制了它们从苍蝇到人类的共同进化。我们的发现不仅是 建立果蝇作为相关的遗传模型来研究HAP40的生理作用，也导致了我们的 假设HAP40是HTT的保守中央调节因子，并可能是突变的关键调节器 HTT毒性。利用在果蝇和培养的哺乳动物细胞中建立的检测和HD模型，我们将 系统地检验这一假设。在目标1，我们将对dhap40进行全面的表型分析。 基因，并检测其与dhtt的遗传交互作用，以获得首次系统评价HAP40在 并在全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亚细胞动力学及其不同细胞功能的中央调节因子，因此 从分子和细胞水平阐明其与HTT的关系。在目标3中，利用油井- 在果蝇和哺乳动物神经元中建立HD模型，我们将严格询问HAP40的作用 突变的HTT毒性。从这些多学科的研究中，我们将首次对 HAP40的生理功能、对内源性HTT功能的影响及HD发病机制。这个 结果为HAP40治疗HD的新途径奠定了基础。

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.