O'Neill Laura P, Spotswood Hugh T, Fernando Milan, Turner Bryan M
Chromatin and Gene Expression Group, Institute of Biomedical Research, University of Birmingham Medical School, Birmingham B15 2TT, UK.
Biol Chem. 2008 Apr;389(4):365-70. doi: 10.1515/BC.2008.046.
Silencing of genes on one of the two female X chromosomes early in development helps balance expression of X-linked genes between XX females and XY males and involves chromosome-wide changes in histone variants and modifications. Mouse female embryonic stem (ES) cells have two active Xs, one of which is silenced on differentiation, and provide a powerful model for studying the dynamics of X inactivation. Here, we use immunofluorescence microscopy of metaphase chromosomes to study changes in H3 mono-, di- or tri-methylated at lysine 4 (H3K4mel, -2 or -3) on the inactivating X (Xi) in female ES cells. H3K4me3 is absent from Xi in approximately 25% of chromosome spreads by day 2 of differentiation and in 40-50% of spreads by days 4-6, making it one of the earliest detectable changes on Xi. In contrast, loss of H3K4me2 occurs 1-2 days later, when histone acetylation also diminishes. Remarkably, H3K4mel is depleted on both (active) X chromosomes in undifferentiated female ES cells, and on the single X in males, and remains depleted on Xi. Consistent with this, chromatin immunoprecipitation reveals differentiation-related reductions in H3K4me2 and H3K4me3 at the promoter regions of genes undergoing X-inactivation in female ES cells, but no comparable change in H3K4me1.
在发育早期使两条雌性X染色体之一上的基因沉默，有助于平衡XX雌性和XY雄性之间X连锁基因的表达，并且涉及组蛋白变体和修饰在全染色体范围的变化。小鼠雌性胚胎干细胞（ES细胞）有两条活跃的X染色体，其中一条在分化时会沉默，这为研究X染色体失活的动态过程提供了一个强大的模型。在这里，我们利用中期染色体的免疫荧光显微镜技术，研究雌性ES细胞中失活X染色体（Xi）上赖氨酸4位点单甲基化、双甲基化或三甲基化的H3（H3K4me1、-2或-3）的变化。在分化第2天时，约25%的染色体铺片中Xi上不存在H3K4me3，到第4 - 6天时，这一比例为40 - 50%，这使其成为Xi上最早可检测到的变化之一。相比之下，H3K4me2的缺失在1 - 2天后出现，此时组蛋白乙酰化也减少。值得注意的是，未分化的雌性ES细胞中两条（活跃的）X染色体以及雄性的单条X染色体上的H3K4me1均缺失，并且在Xi上也一直保持缺失状态。与此一致的是，染色质免疫沉淀显示，雌性ES细胞中经历X染色体失活的基因启动子区域的H3K4me2和H3K4me3与分化相关减少，但H3K4me1没有类似变化。
