Sequence-directed base mispairing in human oncogenes.
Sequence-directed base mispairing in human oncogenes.
复制标题
人类癌基因中序列定向的碱基错配。
DOI：
10.1128/mcb.18.8.4659
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发表时间：
1998
影响因子：
5.3
通讯作者：
Davidson,RL
中科院分区：
文献类型：
--
作者：
Lall,L;Davidson,RL
在人类实体瘤中最常见的癌基因突变是密码子12的GG双联体的3′ G残基发生GC→AT转换。我们先前已经证明，胸苷诱变在哺乳动物细胞中发生具有相同的序列特异性，因为诱变优先发生在GG双联体的3′ G。在这项研究中，在体外DNA合成实验进行评估的影响，本地DNA序列的碱基错配，以确定序列定向突变的机制，胸苷及其可能的关系，激活点突变的N-，Ki-和Ha-ras癌基因在实体瘤。为了避免由于发生错配修复以及碱基错误掺入而使结果解释复杂化，在无修复环境中使用无核酸外切酶的Klenow聚合酶进行实验。这些实验的结果表明，脱氧核糖基胸腺嘧啶（dT）的错误掺入发生在人癌基因密码子12的GG双联体的3′ G相对于5′ G的效率高几倍。这些结果进一步表明，不同癌基因第12位密码子GG双联体3′ G和5′ G对端dT错误掺入程度的相对差异受双联体上游碱基的影响。在GG双联体中，可以看到5′ G和3′ G残基对dT错误掺入的程度具有相互相反的影响。5′ G对dT错误掺入有促进作用，而3′ G对dT错误掺入有抑制作用。据推测，GG双联体内的这些相互作用是加性的，因此在GG双联体中的3′ G和5′ G残基之间观察到的dT错误掺入的巨大差异是这些双联体内组合的刺激和抑制作用的最终结果。由于观察到的GG双联体中dT错误掺入的模式对应于人实体瘤中癌基因激活的最常见模式，这些实验的结果表明，序列指导的dT错误掺入可能参与了人癌基因的激活模式，通过在这些癌基因的密码子12中的GG双联体的3′ G处优先引起GC→AT转换。
The most frequently observed mutations inrasoncogenes in solid human tumors are GC→AT transitions at the 3′ G residue of the GG doublet in codon 12 of these oncogenes. We had shown previously that mutagenesis by thymidine occurred with the same sequence specificity in mammalian cells, in that mutagenesis occurred preferentially at the 3′ G of GG doublets. In this study, in vitro DNA synthesis experiments were carried out to assess the effect of local DNA sequence on base mispairing in order to determine the mechanism of sequence-directed mutagenesis by thymidine and its possible relationship to activating point mutations in N-, Ki- and Ha-rasoncogenes in solid human tumors. To avoid complicating the interpretation of the results because of the occurrence of mismatch repair as well as base misincorporation, the experiments were carried out in a repair-free environment with exonuclease-free Klenow polymerase. The results of these experiments showed that misincorporation of deoxyribosylthymine (dT) occurred with several-fold-greater efficiency opposite the 3′ G compared to the 5′ G of the GG doublet in codon 12 of humanrasoncogenes. These results further demonstrated that the relative difference in the extent of dT misincorporation opposite the 3′ G and the 5′ G of GG doublets in codon 12 in the variousrasoncogenes was affected by the base immediately upstream of the doublet. Within the GG doublet, it was seen that the 5′ G and 3′ G residues had an effect on the extent of dT misincorporation opposite each other. The 5′ G was shown to have a stimulatory effect on dT misincorporation opposite the 3′ G, while the 3′ G was shown to have an inhibitory effect on dT misincorporation opposite the 5′ G. Presumably, these mutual interactions within GG doublets are additive, such that the large differential in dT misincorporation observed between the 3′ G and 5′ G residues in GG doublets is the end result of the combined stimulatory and inhibitory effects within these doublets. Since the observed pattern of dT misincorporation within GG doublets corresponds to the most frequent mode of activation ofrasoncogenes in solid human tumors, the results of these experiments suggest that sequence-directed dT misincorporation may be involved in the pattern of activation of humanrasoncogenes, by causing GC→AT transitions preferentially at the 3′ G of the GG doublet in codon 12 of these oncogenes.