Paterno G D, Adra C N, McBurney M W
Mol Cell Biol. 1985 Oct;5(10):2705-12. doi: 10.1128/mcb.5.10.2705-2712.1985.
The embryonal carcinoma cell line, C86S1, carries two X chromosomes, one of which replicates late during S phase of the cell cycle and appears to be genetically inactive. C86S1A1 is a mutant which lacks activity of the X-encoded enzyme, hypoxanthine phosphoribosyltransferase (HPRT). Treatment of C86S1A1 cells with DNA-demethylating agents, such as 5-azacytidine (5AC), resulted in (i) the transient expression in almost all cells of elevated levels of HPRT and three other enzymes encoded by X-linked genes and (ii) the stable expression of HPRT in up to 5 to 20% of surviving cells. Most cells which stably expressed HPRT had two X chromosomes which replicated in early S phase. C86S1A1 cells which had lost the inactive X chromosome did not respond to 5AC. These results suggest that DNA demethylation results in the reactivation of genes on the inactive X chromosome and perhaps in the reactivation of the entire X chromosome. No such reactivation occurred in C86S1A1 cells when the cells were differentiated before exposure to 5AC. Thus, the process of X chromosome inactivation may be a sequential one involving, as a first step, methylation of certain DNA sequences and, as a second step, some other mechanism(s) of transcriptional repression.
胚胎癌细胞系C86S1含有两条X染色体，其中一条在细胞周期的S期晚期进行复制，似乎在基因上是无活性的。C86S1A1是一种缺乏X编码酶次黄嘌呤磷酸核糖转移酶（HPRT）活性的突变体。用DNA去甲基化剂，如5-氮杂胞苷（5AC）处理C86S1A1细胞，结果是：（i）几乎所有细胞中HPRT和其他三种由X连锁基因编码的酶的水平短暂升高；（ii）在多达5%至20%的存活细胞中HPRT稳定表达。大多数稳定表达HPRT的细胞有两条在S期早期复制的X染色体。丢失了无活性X染色体的C86S1A1细胞对5AC无反应。这些结果表明，DNA去甲基化导致无活性X染色体上的基因重新激活，也许还导致整个X染色体的重新激活。当C86S1A1细胞在暴露于5AC之前分化时，未发生这种重新激活。因此，X染色体失活过程可能是一个连续的过程，第一步涉及某些DNA序列的甲基化，第二步涉及一些其他转录抑制机制。
