Roles of yeast DNA polymerases δ and ζ and of Rev1 in the bypass of abasic sites
Roles of yeast DNA polymerases δ and ζ and of Rev1 in the bypass of abasic sites
复制标题
DOI：
10.1101/gad.882301
复制
发表时间：
2001-04-15
影响因子：
10.5
通讯作者：
Prakash, L
中科院分区：
文献类型：
--
作者：
Haracska, L;Unk, I;Prakash, L
脱碱基（AP）位点是DNA中最常见的损伤之一，它们对复制DNA机制的持续合成具有强烈的阻断作用。在这里，我们展示了酵母DNA聚合酶δ和ζ的联合作用对AP位点的有效旁路。在该反应中，Pol delta插入与AP位点相对的A核苷酸，并且Pol zeta随后从插入的核苷酸延伸。与这些观察结果一致，酵母CAN 1（s）基因突变的序列分析表明，A是最常插入AP位点对面的核苷酸。核苷酸C、G和T也被掺入，但频率要低得多。酶如Rev 1和Pol eta可能有助于这些其他核苷酸的插入;然而，Rev 1在AP旁路中的主要作用可能是结构性的。稳态动力学分析表明，Pol zeta在掺入AP位点对面的核苷酸方面效率极低，但它有效地从插入该损伤对面的核苷酸，特别是A延伸。因此，在真核生物中，绕过AP位点需要两种DNA聚合酶的顺序作用，其中延伸步骤仅取决于Pol zeta，但插入步骤可以变化很大，不仅涉及复制型DNA聚合酶Pol delta的主要作用，还涉及各种跨损伤合成聚合酶的不太突出的作用。
Abasic (AP) sites are one of the most frequently formed lesions in DNA, and they present a strong block to continued synthesis by the replicative DNA machinery. Here we show efficient bypass of an AP site by the combined action of yeast DNA polymerases delta and zeta. In this reaction, Pol delta inserts an A nucleotide opposite the AP site, and Pol zeta subsequently extends from the inserted nucleotide. Consistent with these observations, sequence analyses of mutations in the yeast CAN1(s) gene indicate that A is the nucleotide inserted most often opposite AP sites. The nucleotides C, G, and T are also incorporated, but much less frequently. Enzymes such as Rev1 and Pol eta may contribute to the insertion of these other nucleotides; the predominant role of Rev1 in AP bypass, however, is likely to be structural. Steady-state kinetic analyses show that Pol zeta is highly inefficient in incorporating nucleotides opposite the AP site, but it efficiently extends from nucleotides, particularly an A, inserted opposite this lesion. Thus, in eukaryotes, bypass of an AP site requires the sequential action of two DNA polymerases, wherein the extension step depends solely upon Pol zeta, but the insertion step can be quite varied, involving not only the predominant action of the replicative DNA polymerase, Pol delta, but also the less prominent role of various translesion synthesis polymerases.