Praja2 and the regulation of Ataxin-1 clearance in vivo

描述(申请人提供)：脊髓小脑性共济失调1型(SCA1)是一种成人起病的遗传性疾病，导致小脑浦肯野细胞退化，最终在发病10-30年后死亡。患者首先出现轻微的协调和平衡问题(共济失调)。SCA1是由ATXN1基因的CAG重复突变引起的，编码ATXN1蛋白的聚谷氨酰胺(PolyQ)链异常扩张。随着神经退化的进展，大脑的其他区域也参与进来，导致认知障碍以及言语、吞咽和呼吸控制方面的问题。病理学的基本基础是浦肯野细胞正常功能的异常，影响了RNA剪接和基因转录的调节。谷氨酰胺膨胀的ATXN1具有高度的稳定性和更强的抗降解性。此外，ATXN1中S776的磷酸化是一种已知的影响蛋白质水平的翻译后修饰。本研究旨在探讨小脑浦肯野细胞(PC)中ATXN1清除的细胞机制，小脑浦肯野细胞是SCA1的主要病理部位。Praja2是一种E3泛素蛋白连接酶，介导靶蛋白的多泛素化。初步观察表明，体内PC中的Praja2基因敲除会导致ATXN1蛋白水平异常升高。Praja2在PC中高度表达，有趣的是，在SCA1小鼠模型的小脑中显著下调，表明这一途径可能在疾病中受到影响。这项修订后的提案将探索Praja2对PC中ATXN1清除的调节，以及这可能如何依赖于多聚Q扩展和/或S776磷酸化状态。第一个目标将检查体内敲除Praja2是否调节小脑PC中的ATXN1水平。该提案的第二个目的是研究Praja2是否是ATXN1的泛素蛋白连接酶。第三个目标将检查体内PC中Praja2水平的调节是否影响动物模型中的SCA1病理。更好地了解ATXN1清除在体内是如何调控的，可能为开发治疗SCA1的长期目标提供可行的途径。

DESCRIPTION (provided by applicant): Spinocerebellar ataxia type 1 (SCA1) is an adult-onset, inherited disease that leads to degeneration of Purkinje cells of the cerebellum and culminates in death 10-30 years after disease onset. Patients first present with mild problems with coordination and balance (ataxia). SCA1 is caused by a CAG repeat mutation in the ATXN1 gene, encoding the ATXN1 protein with an abnormally expanded polyglutamine (polyQ) tract. As neurodegeneration progresses, other brain regions get involved and contribute to cognitive deficits as well as problems with speech, swallowing, and control of breathing. The fundamental basis of pathology is an aberration in the normal function of Purkinje cells affecting regulation of RNA splicing and gene transcription. Glutamine-expanded ATXN1 is highly stable and more resistant to degradation. Moreover, phosphorylation at S776 in ATXN1 is a post-translational modification known to influence protein levels. Studies are proposed here to examine the cellular mechanisms by which ATXN1 clearance is regulated in cerebellar Purkinje cells (PCs), a primary site of pathology in SCA1. Praja2 is an E3 ubiquitin-protein ligase that mediates polyubiquitination of target proteins. Preliminary observations show that Praja2 knockdown in PCs in vivo results in abnormally elevated levels of ATXN1 protein. Praja2 is highly expressed in PCs and, intriguingly, is significantly downregulated in cerebella of SCA1 mouse models, suggesting this pathway may be affected in disease. This revised proposal will explore Praja2 regulation of ATXN1 clearance in PCs and how that may be dependent on polyQ expansion and/or S776 phosphorylation status. The first aim will examine whether knocking down Praja2 modulates ATXN1 levels in cerebellar PCs in vivo. The second aim in this proposal is to examine whether Praja2 is a ubiquitin-protein ligase to ATXN1. The third aim will examine whether modulating Praja2 levels in PCs in vivo impacts SCA1 pathology in animal models. A better understanding of how ATXN1 clearance is regulated in vivo may provide feasible avenues for the long-term goal of developing a therapy to treat SCA1.