CMG helicase and DNA polymerase e form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication

Lance D. Langston; Dan Zhang; Olga YurievaRoxana E. GeorgescuJeff FinkelsteinNina Y. YaoChiara IndianicMike E. O’Donnell

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CMG helicase and DNA polymerase e form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication

机译：CMG helicase and DNA polymerase e form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication

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DNA replication in eukaryotes is asymmetric, with separate DNA polymerases (Pol) dedicated to bulk synthesis of the leading and lagging strands. Pol α/primase initiates primers on both strands that are extended by Pol e on the leading strand and by Pol δ on the lagging strand. The CMG (Cdc45-MCM-GINS) helicase surrounds the leading strand and is proposed to recruit Pol e for leading-strand synthesis, but to date a direct interaction between CMG and Pol e has not been demonstrated. While purifying CMG helicase overexpressed in yeast, we detected a functional complex between CMG and native Pol e. Using pure CMG and Pol e, we reconstituted a stable 15-subunitCMG–Pol e complex and showed that it is a functional polymerase–helicase on a model replication fork in vitro. On its own, the Pol2 catalytic subunit of Pol e is inefficient in CMG-dependent replication, but addition of the Dpb2 protein subunit of Pol e, known to bind the Psf1 protein subunit of CMG, allows stable synthesis with CMG. Dpb2 does not affect Pol δ function with CMG, and thus we propose that the connection between Dpb2 andCMGhelps to stabilize Pol e on the leading strand as part of a 15-subunit leading-strand holoenzyme we refer to as CMGE. Direct binding between Pol e and CMG provides an explanation for specific targeting of Pol e to the leading strand and provides clear mechanistic evidence for how strand asymmetry is maintained in eukaryotes.

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《Proceedings of the National Academy of Sciences of the United States of America》 |2014年第43期|15390-15395|共6页
作者
Lance D. Langston; Dan Zhang; Olga YurievaRoxana E. GeorgescuJeff FinkelsteinNina Y. YaoChiara IndianicMike E. O’Donnell;
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作者单位

The Rockefeller University;

Manhattan College, Riverdale, NY 10471;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
原文格式 PDF
正文语种英语
中图分类自然科学总论;
关键词
DNA replication; replication fork; helicase; polymerase; CMG;

CMG helicase and DNA polymerase e form a functional 15-subunit holoenzyme for eukaryotic leading-strand DNA replication

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