Active chromatin marks are retained on X chromosomes lacking gene or repeat silencing despite XIST/Xist expression in somatic cell hybrids.
Active chromatin marks are retained on X chromosomes lacking gene or repeat silencing despite XIST/Xist expression in somatic cell hybrids.
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DOI：
10.1371/journal.pone.0010787
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发表时间：
2010-05-24
期刊：
影响因子：
3.7
通讯作者：
Brown CJ
中科院分区：
文献类型：
--
作者：
Thorogood NP;Brown CJ
X 染色体失活发生在哺乳动物发育的早期，导致失活的 X 染色体获得许多异染色质特征。虽然 XIST 在失活过程中发挥着关键作用，但该 ncRNA 的作用方法尚未确定。为了评估异染色质的哪些特征可以通过 XIST RNA 的表达和定位直接招募，我们分析了小鼠/人体细胞杂交体，其中人和小鼠 XIST/Xist 的表达是通过去甲基化从活性 X 诱导的。此类杂交体先前已被证明能够将 XIST/Xist 表达与基因沉默断开，并且我们确认 X 连锁基因表达的维持，甚至接近 Xist 基因座，尽管小鼠 Xist 进行了局部表达。 XIST/Xist 表达时未观察到活性染色质标记 H3 乙酰化和 H3 赖氨酸 4 甲基化的丢失，也未观察到 DNA 甲基化的增加；因此，兼性异染色质的这些标记不仅仅取决于 Xist 表达。在 Xist 表达时观察到 Cot-1 孔，即与 Cot-1 探针杂交的耗尽 RNA 区域；然而，这些体细胞中这些频率和强度均降低。在体细胞杂交体中观察到与人类染色体相对应的人类 Cot-1 转录域。 X 的 Cot-1 结构域不会随着 XIST 的表达而减少，因此在小鼠体细胞背景中无法定位到人类 X 染色体。尽管存在 X 连锁基因沉默，但小鼠/人类杂交细胞中的人类无活性 X 也显示出离域的 XIST 表达和持续的 Cot-1 结构域。这些结果与最近将 Cot-1 沉默与基因沉默分开的报告一致，但也证明了这些杂交细胞中沉默的 X 染色体的重复元件表达。
X-chromosome inactivation occurs early in mammalian development and results in the inactive X chromosome acquiring numerous hallmarks of heterochromatin. While XIST is a key player in the inactivation process, the method of action of this ncRNA is yet to be determined. To assess which features of heterochromatin may be directly recruited by the expression and localization of the XIST RNA we have analyzed a mouse/human somatic cell hybrid in which expression of human and mouse XIST/Xist has been induced from the active X by demethylation. Such hybrids had previously been demonstrated to disconnect XIST/Xist expression from gene silencing and we confirm maintenance of X-linked gene expression, even close to the Xist locus, despite the localized expression of mouse Xist. Loss of the active chromatin marks H3 acetylation and H3 lysine 4 methylation was not observed upon XIST/Xist expression, nor was there a gain of DNA methylation; thus these marks of facultative heterochromatin are not solely dependent upon Xist expression. Cot-1 holes, regions of depleted RNA hybridization with a Cot-1 probe, were observed upon Xist expression; however, these were at reduced frequency and intensity in these somatic cells. Domains of human Cot-1 transcription were observed corresponding to the human chromosomes in the somatic cell hybrids. The Cot-1 domain of the X was not reduced with the expression of XIST, which fails to localize to the human X chromosome in a mouse somatic cell background. The human inactive X in a mouse/human hybrid cell also shows delocalized XIST expression and an ongoing Cot-1 domain, despite X-linked gene silencing. These results are consistent with recent reports separating Cot-1 silencing from genic silencing, but also demonstrate repetitive element expression from an otherwise silent X chromosome in these hybrid cells.
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