酿酒酵母DNA复制的精确时空控制
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摘要
DNA复制是最基本的生命活动之一。DNA复制本身的错误及其过程控制的异常是细胞内基因组不稳定的主要来源,会导致细胞生长异常、衰老、癌变乃至死亡。为了保证基因组DNA能够精确且完整的复制,DNA复制受到严格的调控。在G1期,DNA复制解旋酶的核心组分Mcm2-7复合体被招募到复制起点,获得复制许可资格。进入S期后,在两个周期性蛋白激酶及多个支架蛋白的作用下,复制解旋酶的激活因子Cdc45和GINS复合体被招募至Mcm2-7,形成解旋酶全酶Cdc45-Mcm2-7-GINS (CMG)复合体。随后,众多复制相关蛋白在精准的时空控制下被招募至CMG平台并组装成复制机器,起始DNA双向复制。当相向而行的两个复制叉相遇,复制机器会从DNA链上解离下来,从而完成DNA复制。关于DNA复制过程的研究在近十年来取得了跨越式的突破。本文以酿酒酵母为例,围绕所有真核生物中都高度保守的DNA复制控制开关——CMG解旋酶,对真核生物DNA复制的最新进展进行综述。
DNA replication is one of the fundamental processes of all lives. Errors produced by DNAreplication and abnormalities in its regulation are major sources of genomic instability in cells, leading toabnormal cell growth, aging, tumorigenesis and even death. In order to ensure accurate and completeduplication of genomic DNA, DNA replication is strictly regulated in all eukaryotes. In the G1 phase, thecore component of the DNA replicative helicase——Mcm2-7 is recruited to the origins. This is calledreplication licensing. After cells entering the S phase, the helicase co-activators Cdc45 and GINS arerecruited to Mcm2-7, forming the helicase holoenzyme Cdc45-Mcm2-7-GINS(CMG) complex. Subsequently,numerous replicating proteins are recruited to the CMG platform under precise spatiotemporal control andassembled into a replication machine to initiate bidirectional replication. When two converging replicationforks encounter each other, CMGs are displaced to terminate the progression of these forks. The last decadehas witnessed the leap-forward breakthroughs in this field, particularly in the model organismSaccharomyces cerevisiae. Here, we summarize the recent advances in eukaryotic DNA replication with afocus on the motor of replisome, DNA helicase CMG.
DNA replication is one of the fundamental processes of all lives. Errors produced by DNAreplication and abnormalities in its regulation are major sources of genomic instability in cells, leading toabnormal cell growth, aging, tumorigenesis and even death. In order to ensure accurate and completeduplication of genomic DNA, DNA replication is strictly regulated in all eukaryotes. In the G1 phase, thecore component of the DNA replicative helicase——Mcm2-7 is recruited to the origins. This is calledreplication licensing. After cells entering the S phase, the helicase co-activators Cdc45 and GINS arerecruited to Mcm2-7, forming the helicase holoenzyme Cdc45-Mcm2-7-GINS(CMG) complex. Subsequently,numerous replicating proteins are recruited to the CMG platform under precise spatiotemporal control andassembled into a replication machine to initiate bidirectional replication. When two converging replicationforks encounter each other, CMGs are displaced to terminate the progression of these forks. The last decadehas witnessed the leap-forward breakthroughs in this field, particularly in the model organismSaccharomyces cerevisiae. Here, we summarize the recent advances in eukaryotic DNA replication with afocus on the motor of replisome, DNA helicase CMG.
引文
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[2]Wu LH,Liu Y,Kong DC.Mechanism of chromosomal DNAreplication initiation and replication fork stabilization in eukaryotes[J].Science China Life Sciences,2014,57(5):482-487吴丽虹,刘阳,孔道春.真核细胞染色体DNA复制起始及复制叉稳定性维持的机制[J].中国科学:生命科学,2013,48(10):824-832
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[4]Li NN,Zhai YL,Zhang YX,et al.Structure of the eukaryotic MCM complex at 3.8?[J].Nature,2015,524(7564):186-191
[5]Evrin C,Clarke P,Zech J,et al.A double-hexameric MCM2-7complex is loaded onto origin DNA during licensing of eukaryotic DNA replication[J].Proceedings of the National Academy of Sciences of the United States of America,2009,106(48):20240-20245
[6]Remus D,Beuron F,Tolun G,et al.Concerted loading of Mcm2-7double hexamers around DNA during DNA replication origin licensing[J].Cell,2009,139(4):719-730
[7]Bell SP,Labib K.Chromosome duplication in Saccharomyces cerevisiae[J].Genetics,2016,203(3):1027-1067
[8]Leonard AC,Méchali M.DNA replication origins[J].Cold Spring Harbor Perspectives in Biology,2013,5(10):a010116
[9]Sun JC,Evrin C,Samel SA,et al.Cryo-EM structure of a helicase loading intermediate containing ORC-Cdc6-Cdt1-MCM2-7bound to DNA[J].Nature Structural&Molecular Biology,2013,20(8):944-951
[10]Yuan ZN,Riera A,Bai L,et al.Structural basis of Mcm2-7replicative helicase loading by ORC-Cdc6 and Cdt1[J].Nature Structural&Molecular Biology,2017,24(3):316-324
[11]Ticau S,Friedman LJ,Ivica NA,et al.Single-molecule studies of origin licensing reveal mechanisms ensuring bidirectional helicase loading[J].Cell,2015,161(3):513-525
[12]Ticau S,Friedman LJ,Champasa K,et al.Mechanism and timing of Mcm2-7 ring closure during DNA replication origin licensing[J].Nature Structural&Molecular Biology,2017,24(3):309-315
[13]Frigola J,He J,Kinkelin K,et al.Cdt1 stabilizes an open MCMring for helicase loading[J].Nature Communications,2017,8:15720
[14]Zhai YL,Li NN,Jiang HL,et al.Unique roles of the non-identical MCM subunits in DNA replication licensing[J].Molecular Cell,2017,67(2):168-179
[15]Frigola J,Remus D,Mehanna A,et al.ATPase-dependent quality control of DNA replication origin licensing[J].Nature,2013,495(7441):339-343
[16]Coster G,Diffley JFX.Bidirectional eukaryotic DNA replication is established by quasi-symmetrical helicase loading[J].Science,2017,357(6348):314-318
[17]Fang DQ,Cao QH,Lou HQ.Sld3-MCM interaction facilitated by Dbf4-dependent kinase defines an essential step in eukaryotic DNA replication initiation[J].Frontiers in Microbiology,2016,7:885
[18]Deegan TD,Yeeles JT,Diffley JF.Phosphopeptide binding by Sld3 links Dbf4-dependent kinase to MCM replicative helicase activation[J].The EMBO Journal,2016,35(9):961-973
[19]Muramatsu S,Hirai K,Tak YS,et al.CDK-dependent complex formation between replication proteins Dpb11,Sld2,Polε,and GINS in budding yeast[J].Genes&Development,2010,24(6):602-612
[20]Tanaka S,Araki H.Helicase activation and establishment of replication forks at chromosomal origins of replication[J].Cold Spring Harbor Perspectives in Biology,2013,5(12):a010371
[21]Watase G,Takisawa H,Kanemaki MT.Mcm10 plays a role in functioning of the eukaryotic replicative DNA helicase,Cdc45-Mcm-GINS[J].Current Biology,2012,22(4):343-349
[22]van Deursen F,Sengupta S,de Piccoli G,et al.Mcm10 associates with the loaded DNA helicase at replication origins and defines a novel step in its activation[J].The EMBO Journal,2012,31(9):2195-2206
[23]Kanke M,Kodama Y,Takahashi TS,et al.Mcm10 plays an essential role in origin DNA unwinding after loading of the CMGcomponents[J].The EMBO Journal,2012,31(9):2182-2194
[24]Yeeles JTP,Deegan TD,Janska A,et al.Regulated eukaryotic DNA replication origin firing with purified proteins[J].Nature,2015,519(7544):431-435
[25]Douglas ME,Diffley JFX.Recruitment of Mcm10 to sites of replication initiation requires direct binding to the minichromosome maintenance(MCM)complex[J].The Journal of Biological Chemistry,2015,291(11):5879-5888
[26]Quan Y,Xia YS,Liu L,et al.Cell-cycle-regulated interaction between Mcm10 and double hexameric Mcm2-7 is required for helicase splitting and activation during S phase[J].Cell Reports,2015,13(11):2576-2586
[27]Douglas ME,Ali FA,Costa A,et al.The mechanism of eukaryotic CMG helicase activation[J].Nature,2018,555(7695):265-268
[28]Fang DQ,Lengronne A,Shi D,et al.Dbf4 recruitment by forkhead transcription factors defines an upstream rate-limiting step in determining origin firing timing[J].Genes&Development,2018.DOI:10.1101/gad.306571.117
[29]Natsume T,Müller CA,Katou Y,et al.Kinetochores coordinate pericentromeric cohesion and early DNA replication by Cdc7-Dbf4 kinase recruitment[J].Molecular Cell,2013,50(5):661-674
[30]Sengupta S,van Deursen F,de Piccoli G,et al.Dpb2 integrates the leading-strand DNA polymerase into the eukaryotic replisome[J].Current Biology,2013,23(7):543-552
[31]de Piccoli G,Katou Y,Itoh T,et al.Replisome stability at defective DNA replication forks is independent of S phase checkpoint kinases[J].Molecular Cell,2012,45(5):696-704
[32]Gambus A,van Deursen F,Polychronopoulos D,et al.A key role for Ctf4 in coupling the MCM2-7 helicase to DNA polymeraseαwithin the eukaryotic replisome[J].The EMBO Journal,2009,28(19):2992-3004
[33]Randell JCW,Bowers JL,Rodríguez HK,et al.Sequential ATPhydrolysis by Cdc6 and ORC directs loading of the Mcm2-7helicase[J].Molecular Cell,2006,21(1):29-39
[34]Morohashi H,Maculins T,Labib K.The amino-terminal TPRdomain of Dia2 tethers SCFDia2 to the replisome progression complex[J].Current Biology,2009,19(22):1943-1949
[35]Yao NY,O’Donnell M.The RFC clamp loader:structure and function[A]//MacNeill S.The Eukaryotic Replisome:a Guide to Protein Structure and Function[M].Dordrecht:Springer,2012,62:259-279
[36]Pellegrini L.The Polα-primase complex[A]//MacNeill S.The Eukaryotic Replisome:a Guide to Protein Structure and Function[M].Dordrecht:Springer,2012,62:157-169
[37]Perera RL,Torella R,Klinge S,et al.Mechanism for priming DNA synthesis by yeast DNA polymeraseα[J].eLife,2013,2:e00482
[38]Burgers PMJ,Kunkel TA.Eukaryotic DNA replication fork[J].Annual Review of Biochemistry,2017,86:417-438
[39]Smith DJ,Whitehouse I.Intrinsic coupling of lagging-strand synthesis to chromatin assembly[J].Nature,2012,483(7390):434-438
[40]McGuffee SR,Smith DJ,Whitehouse I.Quantitative,genome-wide analysis of eukaryotic replication initiation and termination[J].Molecular Cell,2013,50(1):123-135
[41]Dewar JM,Walter JC.Mechanisms of DNA replication termination[J].Nature Reviews Molecular Cell Biology,2017,18(8):507-516
[42]Dewar JM,Budzowska M,Walter JC.The mechanism of DNAreplication termination in vertebrates[J].Nature,2015,525(7569):345-350
[43]Xue HJ,Xu X,Fu YV.New insights in pre-replication complex formation with single-molecule visualization[J].Science Bulletin,2015,60(12):1133-1135
[44]Liu XJ,Lou HQ.Single molecular biology:coming of age in DNA replication[J].Hereditas,2017,39(9):771-774(in Chinese)刘晓晶,楼慧强.DNA复制研究步入单分子时代[J].遗传,2017,39(9):771-774
[45]Xia P,Dou Z,Yao XB.Progress of super-resolution microscopy[J].Chemistry of Life,2015,35(3):430-437(in Chinese)夏鹏,窦震,姚雪彪.超高分辨率显微技术研究进展[J].生命的化学,2015,35(3):430-437
[46]Riera A,Barbon M,Noguchi Y,et al.From structure to mechanism-understanding initiation of DNA replication[J].Genes&Development,2017,31(11):1073-1088
[47]Abid Ali F,Douglas ME,Locke J,et al.Cryo-EM structure of a licensed DNA replication origin[J].Nature Communications,2017,8(1):2241
[48]Georgescu R,Yuan ZN,Bai L,et al.Structure of eukaryotic CMGhelicase at a replication fork and implications to replisome architecture and origin initiation[J].Proceedings of the National Academy of Sciences of the United States of America,2017,114(5):E697-E706
[2]Wu LH,Liu Y,Kong DC.Mechanism of chromosomal DNAreplication initiation and replication fork stabilization in eukaryotes[J].Science China Life Sciences,2014,57(5):482-487吴丽虹,刘阳,孔道春.真核细胞染色体DNA复制起始及复制叉稳定性维持的机制[J].中国科学:生命科学,2013,48(10):824-832
[3]Zou YL,Li LL,Lou HQ.The critical roles of DNA polymerases in genome stability and related human diseases[J].Chinese Bulletin of Life Sciences,2014,26(11):1166-1171(in Chinese)邹友龙,李丽莉,楼慧强.DNA聚合酶在维持基因组稳定性中的多重功能及其相关疾病[J].生命科学,2014,26(11):1166-1171
[4]Li NN,Zhai YL,Zhang YX,et al.Structure of the eukaryotic MCM complex at 3.8?[J].Nature,2015,524(7564):186-191
[5]Evrin C,Clarke P,Zech J,et al.A double-hexameric MCM2-7complex is loaded onto origin DNA during licensing of eukaryotic DNA replication[J].Proceedings of the National Academy of Sciences of the United States of America,2009,106(48):20240-20245
[6]Remus D,Beuron F,Tolun G,et al.Concerted loading of Mcm2-7double hexamers around DNA during DNA replication origin licensing[J].Cell,2009,139(4):719-730
[7]Bell SP,Labib K.Chromosome duplication in Saccharomyces cerevisiae[J].Genetics,2016,203(3):1027-1067
[8]Leonard AC,Méchali M.DNA replication origins[J].Cold Spring Harbor Perspectives in Biology,2013,5(10):a010116
[9]Sun JC,Evrin C,Samel SA,et al.Cryo-EM structure of a helicase loading intermediate containing ORC-Cdc6-Cdt1-MCM2-7bound to DNA[J].Nature Structural&Molecular Biology,2013,20(8):944-951
[10]Yuan ZN,Riera A,Bai L,et al.Structural basis of Mcm2-7replicative helicase loading by ORC-Cdc6 and Cdt1[J].Nature Structural&Molecular Biology,2017,24(3):316-324
[11]Ticau S,Friedman LJ,Ivica NA,et al.Single-molecule studies of origin licensing reveal mechanisms ensuring bidirectional helicase loading[J].Cell,2015,161(3):513-525
[12]Ticau S,Friedman LJ,Champasa K,et al.Mechanism and timing of Mcm2-7 ring closure during DNA replication origin licensing[J].Nature Structural&Molecular Biology,2017,24(3):309-315
[13]Frigola J,He J,Kinkelin K,et al.Cdt1 stabilizes an open MCMring for helicase loading[J].Nature Communications,2017,8:15720
[14]Zhai YL,Li NN,Jiang HL,et al.Unique roles of the non-identical MCM subunits in DNA replication licensing[J].Molecular Cell,2017,67(2):168-179
[15]Frigola J,Remus D,Mehanna A,et al.ATPase-dependent quality control of DNA replication origin licensing[J].Nature,2013,495(7441):339-343
[16]Coster G,Diffley JFX.Bidirectional eukaryotic DNA replication is established by quasi-symmetrical helicase loading[J].Science,2017,357(6348):314-318
[17]Fang DQ,Cao QH,Lou HQ.Sld3-MCM interaction facilitated by Dbf4-dependent kinase defines an essential step in eukaryotic DNA replication initiation[J].Frontiers in Microbiology,2016,7:885
[18]Deegan TD,Yeeles JT,Diffley JF.Phosphopeptide binding by Sld3 links Dbf4-dependent kinase to MCM replicative helicase activation[J].The EMBO Journal,2016,35(9):961-973
[19]Muramatsu S,Hirai K,Tak YS,et al.CDK-dependent complex formation between replication proteins Dpb11,Sld2,Polε,and GINS in budding yeast[J].Genes&Development,2010,24(6):602-612
[20]Tanaka S,Araki H.Helicase activation and establishment of replication forks at chromosomal origins of replication[J].Cold Spring Harbor Perspectives in Biology,2013,5(12):a010371
[21]Watase G,Takisawa H,Kanemaki MT.Mcm10 plays a role in functioning of the eukaryotic replicative DNA helicase,Cdc45-Mcm-GINS[J].Current Biology,2012,22(4):343-349
[22]van Deursen F,Sengupta S,de Piccoli G,et al.Mcm10 associates with the loaded DNA helicase at replication origins and defines a novel step in its activation[J].The EMBO Journal,2012,31(9):2195-2206
[23]Kanke M,Kodama Y,Takahashi TS,et al.Mcm10 plays an essential role in origin DNA unwinding after loading of the CMGcomponents[J].The EMBO Journal,2012,31(9):2182-2194
[24]Yeeles JTP,Deegan TD,Janska A,et al.Regulated eukaryotic DNA replication origin firing with purified proteins[J].Nature,2015,519(7544):431-435
[25]Douglas ME,Diffley JFX.Recruitment of Mcm10 to sites of replication initiation requires direct binding to the minichromosome maintenance(MCM)complex[J].The Journal of Biological Chemistry,2015,291(11):5879-5888
[26]Quan Y,Xia YS,Liu L,et al.Cell-cycle-regulated interaction between Mcm10 and double hexameric Mcm2-7 is required for helicase splitting and activation during S phase[J].Cell Reports,2015,13(11):2576-2586
[27]Douglas ME,Ali FA,Costa A,et al.The mechanism of eukaryotic CMG helicase activation[J].Nature,2018,555(7695):265-268
[28]Fang DQ,Lengronne A,Shi D,et al.Dbf4 recruitment by forkhead transcription factors defines an upstream rate-limiting step in determining origin firing timing[J].Genes&Development,2018.DOI:10.1101/gad.306571.117
[29]Natsume T,Müller CA,Katou Y,et al.Kinetochores coordinate pericentromeric cohesion and early DNA replication by Cdc7-Dbf4 kinase recruitment[J].Molecular Cell,2013,50(5):661-674
[30]Sengupta S,van Deursen F,de Piccoli G,et al.Dpb2 integrates the leading-strand DNA polymerase into the eukaryotic replisome[J].Current Biology,2013,23(7):543-552
[31]de Piccoli G,Katou Y,Itoh T,et al.Replisome stability at defective DNA replication forks is independent of S phase checkpoint kinases[J].Molecular Cell,2012,45(5):696-704
[32]Gambus A,van Deursen F,Polychronopoulos D,et al.A key role for Ctf4 in coupling the MCM2-7 helicase to DNA polymeraseαwithin the eukaryotic replisome[J].The EMBO Journal,2009,28(19):2992-3004
[33]Randell JCW,Bowers JL,Rodríguez HK,et al.Sequential ATPhydrolysis by Cdc6 and ORC directs loading of the Mcm2-7helicase[J].Molecular Cell,2006,21(1):29-39
[34]Morohashi H,Maculins T,Labib K.The amino-terminal TPRdomain of Dia2 tethers SCFDia2 to the replisome progression complex[J].Current Biology,2009,19(22):1943-1949
[35]Yao NY,O’Donnell M.The RFC clamp loader:structure and function[A]//MacNeill S.The Eukaryotic Replisome:a Guide to Protein Structure and Function[M].Dordrecht:Springer,2012,62:259-279
[36]Pellegrini L.The Polα-primase complex[A]//MacNeill S.The Eukaryotic Replisome:a Guide to Protein Structure and Function[M].Dordrecht:Springer,2012,62:157-169
[37]Perera RL,Torella R,Klinge S,et al.Mechanism for priming DNA synthesis by yeast DNA polymeraseα[J].eLife,2013,2:e00482
[38]Burgers PMJ,Kunkel TA.Eukaryotic DNA replication fork[J].Annual Review of Biochemistry,2017,86:417-438
[39]Smith DJ,Whitehouse I.Intrinsic coupling of lagging-strand synthesis to chromatin assembly[J].Nature,2012,483(7390):434-438
[40]McGuffee SR,Smith DJ,Whitehouse I.Quantitative,genome-wide analysis of eukaryotic replication initiation and termination[J].Molecular Cell,2013,50(1):123-135
[41]Dewar JM,Walter JC.Mechanisms of DNA replication termination[J].Nature Reviews Molecular Cell Biology,2017,18(8):507-516
[42]Dewar JM,Budzowska M,Walter JC.The mechanism of DNAreplication termination in vertebrates[J].Nature,2015,525(7569):345-350
[43]Xue HJ,Xu X,Fu YV.New insights in pre-replication complex formation with single-molecule visualization[J].Science Bulletin,2015,60(12):1133-1135
[44]Liu XJ,Lou HQ.Single molecular biology:coming of age in DNA replication[J].Hereditas,2017,39(9):771-774(in Chinese)刘晓晶,楼慧强.DNA复制研究步入单分子时代[J].遗传,2017,39(9):771-774
[45]Xia P,Dou Z,Yao XB.Progress of super-resolution microscopy[J].Chemistry of Life,2015,35(3):430-437(in Chinese)夏鹏,窦震,姚雪彪.超高分辨率显微技术研究进展[J].生命的化学,2015,35(3):430-437
[46]Riera A,Barbon M,Noguchi Y,et al.From structure to mechanism-understanding initiation of DNA replication[J].Genes&Development,2017,31(11):1073-1088
[47]Abid Ali F,Douglas ME,Locke J,et al.Cryo-EM structure of a licensed DNA replication origin[J].Nature Communications,2017,8(1):2241
[48]Georgescu R,Yuan ZN,Bai L,et al.Structure of eukaryotic CMGhelicase at a replication fork and implications to replisome architecture and origin initiation[J].Proceedings of the National Academy of Sciences of the United States of America,2017,114(5):E697-E706