Molecular mechanisms of triplet repeat instability in Huntington's disease

亨廷顿氏病（HD）是一种神经退行性疾病，由脑内CAG重复序列的扩张引起。 亨廷顿蛋白（HTT）基因，导致主要在纹状体和皮质的神经元死亡。CAG重复是 高度不稳定和具有较长遗传CAG重复序列的患者在较早的年龄患上该疾病。重复 道在体细胞中也高度不稳定。CAG的高度年龄依赖性体细胞扩张 在HD患者的纹状体和皮层神经元中观察到重复，但在未受影响的大脑中没有观察到重复 这表明体细胞CAG重复扩增是疾病表现的驱动因素。 较早的疾病发作也与不间断的CAG重复序列的长度和伴随的增加有关。 躯体不稳定这些发现进一步强调了体细胞CAG扩增在疾病中的重要性。 表现。最近在受影响个体中进行的全基因组关联研究揭示了 疾病发病年龄的遗传修饰剂;这些包括错配修复途径的几个基因 (MMR)（MSH 3，MLH 1，PMS 1和PMS 2）以及FAN 1，一种DNA链间交联修复基因。 独立地，在HD小鼠模型中的研究已经揭示了MMR基因Msh 2， Msh 3或Mlh 1降低纹状体中CAG重复的体细胞不稳定性。MMR（标准函数）的作用 其中之一是维持基因组稳定性）进一步得到以下观察结果的支持， 这条途径中的蛋白质识别并处理由DNA的错杂交形成的螺旋外DNA挤出物。 两条含有重复序列的DNA链这些发现支持了这样的观点，即这种挤压的异常MMR 是重复膨胀过程的基础相比之下，在HD小鼠模型中敲除Fan 1会加重CAG。 重复扩张。因此，我们推测，两个相反的DNA修复机制对CAG挤出的作用。 因为MMR促进重复扩增，而FAN 1减弱CAG扩增，所以这些之间的平衡是不确定的。 受影响的神经元细胞中的相反途径可能决定重复扩增的速率， 疾病表现。由于这两个过程的分子细节仍然不清楚，我们的总体研究 目标是整合生物化学，细胞和表型研究，以统一理解 组织/细胞类型特异性CAG重复扩增的机制。在目标1中，我们将比较和对比 MutSβ和FAN 1启动的CAG挤出修复途径的分子特征和不同结果。在 目的2，我们将确定与CAG挤出相关的蛋白质复合物的功能意义。在 目的3：明确PMS 1（MutLβ异源二聚体的一部分）在调节CAG挤出修复中的作用 重复扩张。这些研究的完成不仅有助于阐明CAG重复的机制， 扩大在HD，但也将告知我们的理解的新兴作用，DNA修复体细胞不稳定性 这是其他三联体重复疾病的基础，如1型肌强直性营养不良和脆性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.