Expanded CAG/CTG repeat disorders affect over 1 in 2500 individuals worldwide. Potential therapeutic avenues include gene silencing and modulation of repeat instability. However, there are major mechanistic gaps in our understanding of these processes, which prevent the rational design of an efficient treatment. To address this, we developed a novel system, ParB/ANCHOR-mediated Inducible Targeting (PInT), in which any protein can be recruited at will to a GFP reporter containing an expanded CAG/CTG repeat. Using PInT, we found no evidence that the histone deacetylase HDAC5 or the DNA methyltransferase DNMT1 modulate repeat instability upon targeting to the expanded repeat, suggesting that their effect is independent of local chromatin structure. Unexpectedly, we found that expanded CAG/CTG repeats reduce the effectiveness of gene silencing mediated by HDAC5 or DNMT1 targeting. The repeat-length effect in gene silencing by HDAC5 was abolished by a small molecule inhibitor of HDAC3. Our results have important implications on the design of epigenome editing approaches for expanded CAG/CTG repeat disorders. PInT is a versatile synthetic system to study the effect of any sequence of interest on epigenome editing.

中文翻译：

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扩展的CAG / CTG重复靶向表观基因组编辑介导的抗性基因沉默

扩大的CAG / CTG重复性疾病影响全世界2500多人中的1多人。潜在的治疗途径包括基因沉默和重复不稳定的调节。但是，在我们对这些过程的理解上存在重大的机械缺陷，这阻碍了有效治疗方法的合理设计。为了解决这个问题，我们开发了一种新颖的系统，即ParB / ANCHOR介导的可诱导靶向（PInT），其中任何蛋白质均可随意募集到包含扩展的CAG / CTG重复序列的GFP报告基因中。使用PInT，我们没有发现证据表明组蛋白脱乙酰基酶HDAC5或DNA甲基转移酶DNMT1在靶向扩大的重复序列后会调节重复序列的不稳定性，表明它们的作用与局部染色质结构无关。不料，我们发现，扩大的CAG / CTG重复序列会降低HDAC5或DNMT1靶向介导的基因沉默的有效性。HDAC5的小分子抑制剂消除了HDAC5基因沉默中的重复长度效应。我们的结果对扩展的CAG / CTG重复性疾病表观基因组编辑方法的设计具有重要意义。PInT是一种通用的合成系统，用于研究任何感兴趣序列对表观基因组编辑的影响。