Defining the role of SUMOylation in spinobulbar muscular atrophy

描述(由申请人提供)： 脊髓型肌萎缩症(SBMA)是一种由雄激素受体(AR)基因CAG/谷氨酰胺扩张引起的下运动神经元退行性疾病。突变蛋白(PolyQ AR)经历激素依赖的去折叠和寡聚化，这些步骤对毒性和男性进行性近端肢体和球肌肉无力的发展至关重要。最近的研究表明，由配体引发的AR的翻译后修饰调节毒性。具体地说，多聚Q AR与SUMO(小泛素样修饰物)的结合已被证明削弱了突变蛋白的配体依赖的寡聚。然而，这种情况发生的机制以及它在体内改变疾病表型的程度目前尚不清楚。这项应用的目的是确定多聚Q AR的SUMO化如何影响SBMA的发病。这一建议的中心假设是SUMO化促进了多聚Q AR的降解，从而限制了错误折叠蛋白的积累，降低了毒性。这一假说源于初步数据显示，在SBMA细胞模型中，SUMO化降低了可溶性AR寡聚体的水平。在目标1中，将描述相扑靶向泛素连接酶(STUbL)降解SUMOylated PolyQ AR的程度。这将通过在SBMA的可诱导细胞模型中稳定表达野生型和非SuMOylatable AR突变体，并表征SUMO对STUbL招募、AR泛素化和降解以及细胞毒性的影响来实现。在目标2中，将在SBMA敲入小鼠模型中表征SUMO化的效果。这将通过使用基因打靶来产生表达非SUMOylatable PolyQ AR的SBMA敲入小鼠来实现。然后将进行分析，将运动神经元退化和骨骼肌病理与现有的表达多Q AR的转基因小鼠进行比较，多Q AR是相扑的靶标。这些目标的成功完成有望确定相扑保护作用的潜在机制，并为治疗设计提供相扑通路的特定靶点。

DESCRIPTION (provided by applicant): Spinobulbar muscular atrophy (SBMA) is a degenerative disorder of lower motor neurons caused by a CAG/glutamine expansion in the androgen receptor (AR) gene. The mutant protein (polyQ AR) undergoes hormone-dependent unfolding and oligomerization, steps that are critical to toxicity and to the development of progressive proximal limb and bulbar muscle weakness in men. Recent studies demonstrate that post- translational modifications of the AR triggered by ligand regulate toxicity. Specifically, conjugation of the polyQ AR by SUMO (small ubiquitin-like modifier) has been shown to impair ligand-dependent oligomerization of the mutant protein. However, the mechanism by which this occurs and the extent to which it alters the disease phenotype in vivo are currently unknown. The objective of this application is to determine how SUMOylation of the polyQ AR affects SBMA pathogenesis. The central hypothesis of this proposal is that SUMOylation promotes degradation of the polyQ AR and thereby limits accumulation of the misfolded protein and diminishes toxicity. This hypothesis springs from preliminary data demonstrating that SUMOylation decreases the levels of soluble AR oligomers in cellular models of SBMA. In Aim 1, the extent to which SUMO Targeted Ubiquitin Ligases (STUbLs) degrade the SUMOylated polyQ AR will be delineated. This will be accomplished by stably expressing wild type and non-SUMOylatable AR mutants in an inducible cell model of SBMA and characterizing SUMO-mediated effect on STUbL recruitment, AR ubiquitination and degradation, and cell toxicity. In Aim 2, the effects of SUMOylation will be characterized in a knock-in mouse model of SBMA. This will be accomplished by using gene targeting to generate SBMA knock-in mice that express a non- SUMOylatable polyQ AR. Analyses will then be performed to compare motor neuron degeneration and skeletal muscle pathology with that occurring in existing knock-in mice expressing a polyQ AR that is a target for SUMO. Successful completion of these aims is expected to define the mechanism underlying the protective effects of SUMO and allow for specific targeting of the SUMO pathway for therapeutic design.