Trans-acting factors involved in heterochromatinization of the inactive X chromosome

哺乳动物X染色体失活是一种染色体范围的现象，导致雌性体细胞中两条X染色体中的一条在转录水平上沉默。失活的X染色体具有显著的特性，包括异步DNA复制、X染色体长度的XIST RNA定位、核外围的性染色质小体形成以及组蛋白H4的低乙酰化。然而，这些属性之间的关系及其相对重要性仍然未知。在本研究中，我们试图分离与非活性人类X染色体异染色质有关的反式作用因子。CF150是一种人类单染色体杂交体，其唯一的人类元件是一个或多个非活性人类X染色体(Xi)。在CF150细胞中，导入的xi表现异常：(1)xi没有在细胞核周围形成性染色质小体，(2)xi和其他小鼠染色体一样被适度乙酰化，(3)xi没有被Xist包被。相同的xi被转移到…更多进入HeLa细胞的CF150表现为典型的非活性X，这表明这些特征的充分表现需要某种在CF150中缺失的人类特定因子(S)。通过微细胞融合的方法，通过遗传互补分析确定哪些人类染色体携带相关基因。当从CF150导入到小鼠胚胎癌细胞系中时，XIST的失活发生逆转，但XIST持续表达。尽管X染色体处于活性染色质状态，但被激活的人X染色体上仍包裹着XIST RNA，这些单染色体EC杂交体的细胞分化导致XIST RNA从人X染色体上解离。当同一条重新激活的X染色体从单染色体EC杂交种转移到HeLa细胞中时，重新激活的X染色体被XIST RNA包裹。这些数据共同表明，在小鼠细胞中存在两种不同的Xist RNA定位因素(分别为未分化细胞类型和体细胞类型)。较少

Mammalian X chromosome inactivation is a chromosome-wide phenomenon that leads to transcriptional silencing of one of the two X chromosomes in female somatic cells. The inactivated X chromosome shares striking properties, including asynchronous DNA replication, XIST RNA localization through the length of the X chromosome, sex chromatin body formation at nuclear periphery, and hypoacetylation of histone H4. Relationships among these properties and their relative importance, however, remain unknown. In the present study, we have tried to isolate trans-acting factors involved in heterochromatinization of the inactive human X chromosome.A human monochromosome hybrid, CF150 has one or more inactive human X chromosomes (Xi) as its only human element. In CF150 cells, the introduced Xi behaved abnormally ; (1) Xi did not form sex chromatin body at nucleus peripherally, (2) Xi was moderately acetylated as other mouse chromosomes, and (3) Xi was not coated with XIST RNA.The same Xi transferred f … More rom CF150 into HeLa cells behaved normally as the typical inactive X, suggesting full manifestation of these traits requires a certain human specific factor (s) that is depleted in CF150. A genetic complementation assay has been performed to determine which human chromosome bears the related genes, by microcell fusion.When Xi was introduced from CF150 into mouse embryonal carcinoma (EC) cell lines, the reversal of inactivation occurred, but XIST expressed persistently. Despite of its active chromatin state, the reactivated human X chromosome was coated with XIST RNA.Cell differentiation of these monochromosome EC hybrids leaded the dissociation of XIST RNA from the human X chromosome. When the same reactivated X chromosome was transferred from monochromosome EC hybrids into HeLa cells, the reactivated X chromosome was coated with XIST RNA.Together, these data suggest that two different factors (an undifferentiated and a somatic cell types, respectively) for Xist RNA localization exist in mouse cells. Less