Levine A, Yeivin A, Ben-Asher E, Aloni Y, Razin A
Department of Cellular Biochemistry, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
J Biol Chem. 1993 Oct 15;268(29):21754-9.
The detailed mechanism underlying the inhibition of transcription by DNA methylation is still obscure. Chromatin structure has frequently been proposed as a role player in this mechanism. Histone H1 is a known key element in the formation and stabilization of chromatin fibers. We describe here experiments designed to examine the effect of DNA methylation on the binding of histone H1 to DNA and the consequent inhibitory effect of the bound histone H1 on in vitro transcription. The results of these experiments showed a clear preferential binding of histone H1 to methylated DNA as compared with unmethylated DNA. The in vitro transcription assay indicated that transcription of methylated templates was inhibited at a lower histone H1/DNA ratio than of unmethylated templates, and that the extent of inhibition depends on the density of methyl groups in the promoter region. This inhibition of in vitro transcription was alleviated efficiently by methylated competitor DNA, whereas, under similar conditions, almost no effect was observed with unmethylated competitor. Experiments designed to pinpoint the stage in the transcription process that was suppressed by the preferred binding of histone H1 to methylated template revealed that inhibition occurred at the initiation and not at the elongation level.
DNA甲基化抑制转录的具体机制仍不清楚。染色质结构常被认为是这一机制中的一个作用因素。组蛋白H1是染色质纤维形成和稳定过程中一个已知的关键元件。我们在此描述了旨在研究DNA甲基化对组蛋白H1与DNA结合的影响以及结合的组蛋白H1对体外转录的抑制作用的实验。这些实验结果表明，与未甲基化的DNA相比，组蛋白H1明显更倾向于与甲基化的DNA结合。体外转录试验表明，与未甲基化模板相比，甲基化模板的转录在较低的组蛋白H1/DNA比率下就受到抑制，且抑制程度取决于启动子区域甲基基团的密度。甲基化的竞争DNA能有效缓解这种体外转录抑制，而在类似条件下，未甲基化的竞争DNA几乎没有作用。旨在确定转录过程中因组蛋白H1与甲基化模板的优先结合而受到抑制的阶段的实验表明，抑制发生在起始阶段而非延伸阶段。
