Department of Genetics, Health Sciences Center, Louisiana State University, New Orleans, LA 70112, USA.
J Biol Chem. 2012 Aug 24;287(35):29958-67. doi: 10.1074/jbc.M112.356758. Epub 2012 Jul 11.
While DNA repair has been implicated in CAG·CTG repeat expansion, its role in the GAA·TTC expansion of Friedreich ataxia (FRDA) is less clear. We have developed a human cellular model that recapitulates the DNA repeat expansion found in FRDA patient tissues. In this model, GAA·TTC repeats expand incrementally and continuously. We have previously shown that the expansion rate is linked to transcription within the repeats. Our working hypothesis is that structures formed within the GAA·TTC repeat during transcription attract DNA repair enzymes that then facilitate the expansion process. MutSβ, a heterodimer of MSH2 and MSH3, is known to have a role in CAG·CTG repeat expansion. We now show that shRNA knockdown of either MSH2 or MSH3 slowed GAA·TTC expansion in our system. We further characterized the role of MutSβ in GAA·TTC expansion using a functional assay in primary FRDA patient-derived fibroblasts. These fibroblasts have no known propensity for instability in their native state. Ectopic expression of MSH2 and MSH3 induced GAA·TTC repeat expansion in the native FXN gene. MSH2 is central to mismatch repair and its absence or reduction causes a predisposition to cancer. Thus, despite its essential role in GAA·TTC expansion, MSH2 is not an attractive therapeutic target. The absence or reduction of MSH3 is not strongly associated with cancer predisposition. Accordingly, MSH3 has been suggested as a therapeutic target for CAG·CTG repeat expansion disorders. Our results suggest that MSH3 may also serve as a therapeutic target to slow the expansion of GAA·TTC repeats in the future.
虽然 DNA 修复与 CAG·CTG 重复扩展有关，但它在 Friedreich 共济失调（FRDA）的 GAA·TTC 扩展中的作用尚不清楚。我们开发了一种人类细胞模型，该模型重现了 FRDA 患者组织中发现的 DNA 重复扩展。在该模型中，GAA·TTC 重复不断递增和连续扩展。我们之前已经表明，扩展率与重复内的转录有关。我们的工作假设是，在转录过程中在 GAA·TTC 重复内形成的结构吸引 DNA 修复酶，然后促进扩展过程。MutSβ 是 MSH2 和 MSH3 的异二聚体，已知在 CAG·CTG 重复扩展中具有作用。我们现在表明，shRNA 敲低 MSH2 或 MSH3 均可减缓我们系统中的 GAA·TTC 扩展。我们进一步使用原发性 FRDA 患者衍生的成纤维细胞中的功能测定来表征 MutSβ 在 GAA·TTC 扩展中的作用。这些成纤维细胞在其自然状态下没有不稳定的倾向。MSH2 和 MSH3 的异位表达诱导了 FXN 基因中 GAA·TTC 重复扩展。MSH2 是错配修复的核心，其缺失或减少会导致癌症易感性。因此，尽管 MSH2 在 GAA·TTC 扩展中具有重要作用，但它不是一个有吸引力的治疗靶标。MSH3 的缺失或减少与癌症易感性没有强烈关联。因此，MSH3 被提议作为 CAG·CTG 重复扩展疾病的治疗靶标。我们的结果表明，MSH3 也可能成为未来减缓 GAA·TTC 重复扩展的治疗靶标。
