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首页> 外文期刊>Molecular and Cellular Biology >Methylation-Mediated Proviral Silencing Is Associated with MeCP2 Recruitment and Localized Histone H3 Deacetylation
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Methylation-Mediated Proviral Silencing Is Associated with MeCP2 Recruitment and Localized Histone H3 Deacetylation

机译：甲基化介导的前病毒沉默与MeCP2招聘和局部组蛋白H3脱乙酰化有关

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The majority of 5-methylcytosine in mammalian DNA resides in endogenous transposable elements and is associated with the transcriptional silencing of these parasitic elements. Methylation also plays an important role in the silencing of exogenous retroviruses. One of the difficulties inherent in the study of proviral silencing is that the sites in which proviruses randomly integrate influence the probability of de novo methylation and expression. In order to compare methylated and unmethylated proviruses at the same genomic site, we used a recombinase-based targeting approach to introduce an in vitro methylated or unmethylated Moloney murine leukemia-based provirus in MEL cells. The methylated and unmethylated states are maintained in vivo, with the exception of the initially methylated proviral enhancer, which becomes demethylated in vivo. Although the enhancer is unmethylated and remodeled, the methylated provirus is transcriptionally silent. To further analyze the repressed state, histone acetylation status was determined by chromatin immunoprecipitation (ChIP) analyses, which revealed that localized histone H3 but not histone H4 hyperacetylation is inversely correlated with proviral methylation density. Since members of the methyl-CpG binding domain (MBD) family of proteins recruit histone deacetylase activity, these proteins may play a role in proviral repression. Interestingly, only MBD3 and MeCP2 are expressed in MEL cells. ChIPs with antibodies specific for these proteins revealed that only MeCP2 associates with the provirus in a methylation-dependent manner. Taken together, our results suggest that MeCP2 recruitment to a methylated provirus is sufficient for transcriptional silencing, despite the presence of a remodeled enhancer.
机译：哺乳动物DNA中的大多数5-甲基胞嘧啶位于内源转座因子中，并与这些寄生因子的转录沉默有关。甲基化在外源逆转录病毒的沉默中也起重要作用。前病毒沉默研究固有的困难之一是前病毒随机整合的位点影响从头甲基化和表达的可能性。为了在相同的基因组位点比较甲基化和未甲基化的原病毒，我们使用了基于重组酶的靶向方法在MEL细胞中引入了基于体外甲基化或未甲基化的莫洛尼鼠白血病的原病毒。甲基化和未甲基化状态在体内得以维持，但最初的甲基化原病毒增强剂除外，后者在体内变得脱甲基。尽管增强子是未甲基化和重塑的，但是甲基化的原病毒在转录上是沉默的。为了进一步分析阻抑状态，通过染色质免疫沉淀（ChIP）分析确定组蛋白乙酰化状态，这揭示了局部组蛋白H3而非组蛋白H4过乙酰化与原病毒甲基化密度成反比。由于蛋白质的甲基CpG结合域（MBD）家族成员募集了组蛋白脱乙酰基酶活性，因此这些蛋白质可能在前病毒抑制中起作用。有趣的是，仅MBD3和MeCP2在MEL细胞中表达。具有针对这些蛋白质的特异性抗体的ChIP显示，只有MeCP2以甲基化依赖的方式与原病毒缔合。两者合计，我们的结果表明，尽管存在重塑的增强子，MeCP2募集到甲基化的前病毒足以实现转录沉默。

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