Mathison B H, Said B, Shank R C
Department of Community and Environmental Medicine, University of California, Irvine 92717.
Carcinogenesis. 1993 Feb;14(2):323-7. doi: 10.1093/carcin/14.2.323.
Effects on N-methyl-N-nitrosourea (MNU) mediated methylation of the N7 position of guanine were compared in defined sequences of DNA containing cytosine or 5-methylcytosine (5mC) using a Maxam-Gilbert sequencing technique. Cytosine methylation in 5'-CpG-3' pairs within a subcloned fragment of the 5' region of the human HPRT gene was generated with SssI methylase and S-adenosylmethionine. Cytosine methylation was demonstrated by both the inhibition of DNA restriction by methylation sensitive endonucleases and the lack of cleavage at 5-methylcytosines by hydrazine. MNU-dependent methylation of the N7 position of guanine was inhibited up to 18% when 5mC was a 5' neighboring base to guanine and was inhibited up to 36% in an alternating CpG region in which both 5' and 3' neighboring bases of guanine were enzymatically altered to 5mC. It can be concluded that 5-methylcytosine has discernible effects on MNU methylation of the N7 position of specific guanine bases in DNA.
使用Maxam-Gilbert测序技术，比较了在含有胞嘧啶或5-甲基胞嘧啶（5mC）的特定DNA序列中，N-甲基-N-亚硝基脲（MNU）介导的鸟嘌呤N7位甲基化的影响。使用SssI甲基转移酶和S-腺苷甲硫氨酸在人HPRT基因5'区域亚克隆片段内的5'-CpG-3'对中产生胞嘧啶甲基化。通过甲基化敏感核酸内切酶对DNA限制的抑制以及肼对5-甲基胞嘧啶的切割缺乏来证明胞嘧啶甲基化。当5mC是鸟嘌呤的5'相邻碱基时，鸟嘌呤N7位的MNU依赖性甲基化被抑制高达18%，在交替的CpG区域中，鸟嘌呤的5'和3'相邻碱基都被酶促改变为5mC时，抑制高达36%。可以得出结论，5-甲基胞嘧啶对DNA中特定鸟嘌呤碱基的N7位MNU甲基化有明显影响。
