Hatayama T, Nakamura T, Yukioka M
Biochem Int. 1984 Aug;9(2):251-8.
To examine the distribution of 5-methylcytosine in chromatin DNA, DNA of HeLa cells was labeled with [3H-methyl]methionine and [14C] thymidine and analyzed after extensive digestion of the nuclei with micrococcal nuclease. When the chromatin solubilized with the nuclease was fractionated on a sucrose density gradient, DNA in mononucleosomes was considerably depleted in 5-methylcytosine, as compared with polynucleosomes. Electrophoretic separation of DNA from the chromatin also revealed the depletion of 5-methylcytosine in the mononucleosomal size of DNA. This was confirmed by the chromatographic analysis of 5-methyldeoxycytidine after enzymatic digestion of the DNA to nucleosides. Thus the DNA in mononucleosomes solubilized by extensive micrococcal nuclease digestion is depleted in 5-methylcytosine, suggesting that 5-methylcytosine is preferentially missing from the DNA in the nucleosome core particles.
为了检测染色质DNA中5-甲基胞嘧啶的分布情况，用[³H-甲基]甲硫氨酸和[¹⁴C]胸苷对HeLa细胞的DNA进行标记，并在用微球菌核酸酶对细胞核进行充分消化后进行分析。当用核酸酶溶解的染色质在蔗糖密度梯度上进行分级分离时，与多核小体相比，单核小体中的DNA在5-甲基胞嘧啶方面明显减少。对染色质中的DNA进行电泳分离也显示，在单核小体大小的DNA中5-甲基胞嘧啶减少。在将DNA酶解为核苷后对5-甲基脱氧胞苷进行色谱分析，证实了这一点。因此，通过广泛的微球菌核酸酶消化溶解的单核小体中的DNA在5-甲基胞嘧啶方面减少，这表明核小体核心颗粒中的DNA优先缺失5-甲基胞嘧啶。
