Replication Bypass of N2-Deoxyguanosine Interstrand Cross-Links by Human DNA Polymerases η and ι
Replication Bypass of N2-Deoxyguanosine Interstrand Cross-Links by Human DNA Polymerases η and ι
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DOI：
10.1021/tx300011w
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发表时间：
2012-03-01
影响因子：
4.1
通讯作者：
Minko, Irina G.
中科院分区：
文献类型：
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作者：
Klug, Alex R.;Harbut, Michael B.;Minko, Irina G.
DNA-链间交链(ICL)可以通过需要能够跨损伤DNA合成(TLS)的DNA聚合酶的生化途径来修复。TLS聚合酶在这些途径中的预期功能是在连接位点之外插入相反的核苷酸。这些反应的结果可以是无差错的，也可以是诱变的。脱氧鸟苷(N-2-DG)的外环氮基(N-2-DG)之间形成的ICL的TLS依赖性修复可导致低频率的碱基替换，主要是G到T的颠换。此前，我们在体外证明了由丙烯醛介导的N-2-DG ICL模型的无错误旁路可以由人聚合酶(Polkappa)kappa完成，而Rev1可以通过在交联型DG相反的位置插入DC来实现这种旁路。目前的研究描述了另外两种人类DNA聚合酶，polETA和polIOTA，关于它们对这些病变的无错误或突变旁路的潜在贡献。在单个dNTPs存在的情况下，polETA可以插入dA、dG和dT，但DC的掺入不明显。仅在DC和DG3‘端观察到进一步的引物延伸，且产物量低于匹配的未损伤底物。对加合物外的旁路产物的分析表明，在大多数延伸的引物中，DG与交联型DG相对，并且经常检测到短缺失。当Poliota被测试其复制能力超过这个ICL时，正确的DC优先被掺入，但没有观察到进一步的延伸。在稳态条件下，DC掺入效率比未损伤的DG降低了近500倍。因此，除了Polkappa催化的N-2-DG ICL的无错误旁路之外，可能存在另一种尽管效率较低的机制。在此途径中，Rev1或poliota可以插入与损伤相对的DC，而polETA可以执行后续的延伸。
DNA-interstrand cross-links (ICLs) can be repaired by biochemical pathways requiring DNA polymerases that are capable of translesion DNA synthesis (TLS). The anticipated function of TLS polymerases in these pathways is to insert nucleotides opposite and beyond the linkage site. The outcome of these reactions can be either error-free or mutagenic. TLS-dependent repair of ICLs formed between the exocyclic nitrogens of deoxyguanosines (N-2-dG) can result in low-frequency base substitutions, predominantly G to T transversions. Previously, we demonstrated in vitro that error-free bypass of a model acrolein-mediated N-2-dG ICL can be accomplished by human polymerase (pol) kappa, while Rev1 can contribute to this bypass by inserting dC opposite the crosslinked dG. The current study characterized two additional human DNA polymerases, pol eta and pol iota, with respect to their potential contributions to either error-free or mutagenic bypass of these lesions. In the presence of individual dNTPs, pol eta could insert dA, dG, and dT opposite the cross-linked dG, but incorporation of dC was not apparent. Further primer extension was observed only from the dC and dG 3' termini, and the amounts of products were low relative to the matched undamaged substrate. Analyses of bypass products beyond the adducted site revealed that dG was present opposite the cross-linked dG in the majority of extended primers, and short deletions were frequently detected. When pol iota was tested for its ability to replicate past this ICL, the correct dC was preferentially incorporated, but no further extension was observed. Under the steady-state conditions, the efficiency of dC incorporation was reduced similar to 500-fold relative to the undamaged dG. Thus, in addition to pol kappa-catalyzed error-free bypass of N-2-dG ICLs, an alternative, albeit low-efficiency, mechanism may exist. In this pathway, either Rev1 or pol iota could insert dC opposite the lesion, while pol eta could perform the subsequent extension.