Roles of DNA polymerase I in leading and lagging-strand replication defined by a high-resolution mutation footprint of ColE1 plasmid replication.
Roles of DNA polymerase I in leading and lagging-strand replication defined by a high-resolution mutation footprint of ColE1 plasmid replication.
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DOI：
10.1093/nar/gkr157
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发表时间：
2011-09-01
影响因子：
14.9
通讯作者：
Camps M
中科院分区：
文献类型：
--
作者：
Allen JM;Simcha DM;Ericson NG;Alexander DL;Marquette JT;Van Biber BP;Troll CJ;Karchin R;Bielas JH;Loeb LA;Camps M
DNA聚合酶I（pol I）在滞后链合成期间加工RNA引物，并在DNA修复反应期间填充小间隙。然而，目前还不清楚pol I和pol III在复制和修复过程中如何一起工作，或者pol I在体内对冈崎片段的加工有多广泛。在这里，我们解决这些问题，通过分析易出错的复制ColE 1质粒产生的pol I突变。通过直接测序获得数据，从而可以准确测定突变谱和分布。Pol I的突变足迹表明：（i）在前导链复制过程中，Pol I在至少1.3 kb上逐渐被Pol III取代;（ii）Okazaki片段的Pol I加工仅限于250 nt;（iii）Okazaki片段的大小很短（250 nt）。虽然基于ColE 1质粒复制，但我们的研究结果可能与其他pol I复制过程相关，如染色体复制和DNA修复，这些过程与ColE 1复制主要在招募步骤上不同。这种突变足迹法应该有助于建立其他原核或真核聚合酶在体内的作用，并提供了一种工具，以调查序列拓扑结构，DNA损伤，或与蛋白质的相互作用，可能会影响个别DNA聚合酶的功能。
DNA polymerase I (pol I) processes RNA primers during lagging-strand synthesis and fills small gaps during DNA repair reactions. However, it is unclear how pol I and pol III work together during replication and repair or how extensive pol I processing of Okazaki fragments is in vivo. Here, we address these questions by analyzing pol I mutations generated through error-prone replication of ColE1 plasmids. The data were obtained by direct sequencing, allowing an accurate determination of the mutation spectrum and distribution. Pol I’s mutational footprint suggests: (i) during leading-strand replication pol I is gradually replaced by pol III over at least 1.3 kb; (ii) pol I processing of Okazaki fragments is limited to ∼20 nt and (iii) the size of Okazaki fragments is short (∼250 nt). While based on ColE1 plasmid replication, our findings are likely relevant to other pol I replicative processes such as chromosomal replication and DNA repair, which differ from ColE1 replication mostly at the recruitment steps. This mutation footprinting approach should help establish the role of other prokaryotic or eukaryotic polymerases in vivo, and provides a tool to investigate how sequence topology, DNA damage, or interactions with protein partners may affect the function of individual DNA polymerases.
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DOI：
10.1073/pnas.77.5.2450
发表时间：
1980-01-01
期刊：
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA-BIOLOGICAL SCIENCES
影响因子：
--
作者：
ITOH, T;TOMIZAWA, J
通讯作者：
TOMIZAWA, J
影响因子：
3.2
作者：
KINGSBURY, DT;HELINSKI, DR
通讯作者：
HELINSKI, DR
影响因子：
3.6
作者：
Makiela-Dzbenska, Karolina;Jaszczur, Malgorzata;Fijalkowska, Iwona J.
通讯作者：
Fijalkowska, Iwona J.
影响因子：
5.6
作者：
KITANI, T;YODA, KY;OKAZAKI, T
通讯作者：
OKAZAKI, T
影响因子：
4.8
作者：
Loh, Ern;Choe, Juno;Loeb, Lawrence A.
通讯作者：
Loeb, Lawrence A.