Molecular mechanisms of triplet repeat instability in Huntington's disease

亨廷顿病(HD)是一种神经退行性疾病，由体内CAG重复束扩张引起 亨廷顿蛋白(HTT)基因，导致神经元死亡，主要在纹状体和皮质。CAG重复序列是 高度不稳定和遗传CAG重复较长的患者在较早的年龄患上这种疾病。重复 肌束在体细胞中也是高度不稳定的。慢性萎缩性胃炎高度的年龄依赖性躯体扩张 在HD患者的纹状体和皮质神经元中都可以观察到重复序列，但在未受影响的大脑中没有观察到重复序列 小脑等区域，表明躯体CAG重复扩增是疾病表现的驱动因素。 发病较早也与不间断CAG重复的长度和伴随的增加有关 在躯体不稳定中。这些发现进一步强调了躯体CAG扩张在疾病中的重要性 显化。最近在受影响的个体中进行的全基因组关联研究揭示了存在 疾病发病年龄的遗传修饰物；其中包括错配修复途径的几个基因 (MMR)(MSH3、MLH1、PMS1和PMS2)以及DNA链间交联修复基因FAN1。 独立地，对HD小鼠模型的研究表明，MMR基因Msh2， Msh3或MLH1减少纹状体CAG重复序列的体细胞不稳定性。MMR(规范函数)的角色 其中是为了保持基因组稳定性)在CAG重复扩增中，进一步支持观察到 该途径中的蛋白质识别和处理由错配形成的螺旋外DNA挤出物 两条含有重复序列的DNA链。这些发现支持了这样一种观点，即这种挤出的异常MMR 这是重复扩张过程的基础。相比之下，HD小鼠模型中Fan1基因的敲除加剧了CAG 重复扩展。因此，我们假设两种相反的DNA修复机制作用于CAG挤出。 由于MMR促进重复扩展，而FAN1抑制CAG扩展，因此两者之间的平衡 受影响神经细胞中相反的路径可能决定重复扩张的速度，因此， 疾病表现。由于这两个过程的分子细节仍不清楚，我们最主要的 目标是整合生化、细胞和表型研究，以形成对 组织/细胞类型特异性CAG重复扩增机制。在目标1中，我们将比较和对比 MutSβ和FAN1启动的CAG挤出修复通路的分子特征和不同结果。在……里面 目的2，我们将确定与CAG挤压相关的蛋白质复合体的功能意义。在……里面 目的3，我们将确定pMS1(作为MutLβ异源二聚体的一部分)在CAG挤出修复调节中的作用 然后重复扩张。这些研究的完成不仅将有助于阐明CAG重复的机制 在HD中的扩展，也将使我们了解DNA修复在躯体不稳定中的新角色 这是其他三联体重复疾病的基础，如强直性肌营养不良1型和脆性X相关疾病。

Huntington’s disease (HD) is a neurodegenerative disorder caused by an expansion of a CAG repeat tract within the huntingtin (HTT) gene, leading to neuronal death primarily in the striatum and the cortex. The CAG repeat is highly unstable and patients with longer inherited CAG repeats develop the disease at an earlier age. The repeat tract is also highly unstable in somatic cells. A high degree of age-dependent somatic expansion of the CAG repeat is observed in neurons of both the striatum and the cortex of HD patients, but not in unaffected brain regions like the cerebellum, indicating that somatic CAG repeat expansion is a driver of disease manifestation. Earlier disease onset is also associated with the length of uninterrupted CAG repeats and a concomitant increase in somatic instability. These findings further underscore the importance of somatic CAG expansion in disease manifestation. Recent genome-wide association studies in affected individuals have revealed the existence of genetic modifiers of the age of onset of the disease; these include several genes of the mismatch repair pathway (MMR) (MSH3, MLH1, PMS1, and PMS2) as well as FAN1, a DNA interstrand cross-link repair gene. Independently, studies in mouse models of HD have revealed that genetic knockout of the MMR genes, Msh2, Msh3, or Mlh1 reduces somatic instability of CAG repeats in the striatum. A role for MMR (the canonical function of which is to maintain genomic stability) in CAG repeat expansion is further supported by the observation that proteins in this pathway recognize and process extrahelical DNA extrusions formed by mishybridization of the two repeat-containing DNA strands. These findings support the view that aberrant MMR of such extrusions underlies the repeat expansion process. By contrast, knockout of Fan1 in an HD mouse model exacerbates CAG repeat expansion. We therefore hypothesize that two opposing DNA repair mechanisms act on CAG extrusions. Because MMR promotes repeat expansion, and FAN1 attenuates CAG expansion, the balance between these opposing pathways in affected neuronal cells likely determines the rate of repeat expansion and consequently, disease manifestation. Since the molecular details of either of these processes remains unclear, our overarching goal is to integrate biochemical, cellular, and phenotypic studies to develop a unified understanding of the mechanism of tissue/cell type specific CAG repeat expansion. In Aim 1, we will compare and contrast the molecular features and differing outcomes of the MutSβ- and FAN1- initiated CAG extrusion repair pathways. In Aim 2, we will determine the functional significance of protein complexes that associate with CAG extrusions. In Aim 3, we will define the role of PMS1 (as part of the MutLβ heterodimer) in regulation of CAG extrusion repair and repeat expansion. Completion of these studies will not only shed light on the mechanisms of CAG repeat expansion in HD, but also will inform our understanding of the emerging role of DNA repair in somatic instability that underlies other triplet repeat diseases like myotonic dystrophy type 1 and fragile-X related disorders.