A novel DNA nuclease is stimulated by association with the GINS complex

Li, Zhuo; Pan, Miao; Santangelo, Thomas J.; Chemnitz, Wiebke; Wang, Yuan; Edwards, John A.; Hurwitz, Jerard; Reeve, John N.; Kelman, Zvi

doi:10.1093/nar/gkr181

Cited by 61 publications

(105 citation statements)

References 34 publications

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“…Pol D also interacts strongly with PCNA, conferring the ability to extend rapidly along long stretches of template DNA (28-30). These properties and fast, accurate, and processive DNA synthesis together with evidence that Pol D assembles in vivo into complexes that contain other replisome components (13,31,32) provide consistent support for the hypothesis that Pol D may replicate archaeal genomic DNA.To test this hypothesis, we took advantage of the genetic technologies now available for Thermococcus kodakarensis (33) to manipulate the in vivo structure and expression of Pol B (encoded by TK0001 [34]) (Fig. 2) and Pol D (encoded by TK1902 [DP1] and TK1903 [DP2]).…”

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confidence: 87%

Archaeal DNA Polymerase D but Not DNA Polymerase B Is Required for Genome Replication in Thermococcus kodakarensis

et al. 2013

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c Three evolutionarily distinct families of replicative DNA polymerases, designated polymerase B (Pol B), Pol C, and Pol D, have been identified. Members of the Pol B family are present in all three domains of life, whereas Pol C exists only in Bacteria and Pol D exists only in Archaea. Pol B enzymes replicate eukaryotic chromosomal DNA, and as members of the Pol B family are present in all Archaea, it has been assumed that Pol B enzymes also replicate archaeal genomes. Here we report the construction of Thermococcus kodakarensis strains with mutations that delete or inactivate key functions of Pol B. T. kodakarensis strains lacking Pol B had no detectable loss in viability and no growth defects or changes in spontaneous mutation frequency but had increased sensitivity to UV irradiation. In contrast, we were unable to introduce mutations that inactivated either of the genes encoding the two subunits of Pol D. The results reported establish that Pol D is sufficient for viability and genome replication in T. kodakarensis and argue that Pol D rather than Pol B is likely the replicative DNA polymerase in this archaeon. The majority of Archaea contain Pol D, and, as discussed, if Pol D is the predominant replicative polymerase in Archaea, this profoundly impacts hypotheses for the origin(s), evolution, and distribution of the different DNA replication enzymes and systems now employed in the three domains of life. DNA replication, an essential event for all cellular life, is catalyzed by protein complexes designated replisomes, in which individual activities are tightly regulated and coordinated. DNA polymerases are the functional center of the replisome, but structurally distinct DNA polymerases, designated family C (Pol C) and family B (Pol B) polymerases, catalyze genome replication in Bacteria and eukaryotes, respectively (1-3). This difference has led to much debate, most fundamentally regarding whether DNA replication has evolved more than once, possibly independently in different biological lineages (1, 4-10). All known archaeal genomes encode at least one member of the Pol B family, and given that Archaea are evolutionarily closer to eukaryotes than are Bacteria (11, 12), it has been tacitly assumed, but challenged (13,14), that Pol B enzymes must also replicate archaeal genomes. Presumably, this must be the case for the Crenarchaeota, as their genomes appear to encode only Pol B enzymes. This is, however, only an assumption for all members of the Euryarchaeota, Thaumarchaeota, Korarchaeota, Aigarchaeota, and Nanoarchaeota lineages, as their genomes encode not only Pol B enzymes but also members of an archaeon-specific DNA polymerase family designated Pol D (Fig. 1) (11, 13-15).The Thermococcales are hyperthermophilic Euryarchaea, and given the commercial value of thermostable processive DNA polymerases, Pol B polymerases from this genus have received extensive characterization (13,16,17). Within these single polypeptide enzymes, the regions and residues directly responsible for nucleotide polymerization, 3=¡...

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Archaeal DNA Polymerase D but Not DNA Polymerase B Is Required for Genome Replication in Thermococcus kodakarensis

et al. 2013

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“…Archaea possess a simple homohexameric MCM (5, 7). In addition, archaeal homologs of GINS and Cdc45 have been identified (8)(9)(10)(11)(12)(13)(14). In species of the genus Sulfolobus, we have previously demonstrated that the GINS complex interacts with the N-terminal domains of MCM (8).…”

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confidence: 99%

Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM

Gristwood²,

Hodgson³

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The regulated recruitment of Cdc45 and GINS is key to activating the eukaryotic MCM(2-7) replicative helicase. We demonstrate that the homohexameric archaeal MCM helicase associates with orthologs of GINS and Cdc45 in vivo and in vitro. Association of these factors with MCM robustly stimulates the MCM helicase activity. In contrast to the situation in eukaryotes, archaeal Cdc45 and GINS form an extremely stable complex before binding MCM. Further, the archaeal GINS • Cdc45 complex contains two copies of Cdc45. Our analyses give insight into the function and evolution of the conserved core of the archaeal/eukaryotic replisome.T he initiation of DNA replication is an important control point in the progression of the cell cycle. In eukaryotes, origins are defined by the initiator protein ORC complex that, via the actions of two additional factors, the helicase coloaders Cdc6 and Cdt1, directs loading of the MCM(2-7) replicative helicase onto doublestranded DNA. Activation of the MCM(2-7) replicative helicase occurs subsequent to recruitment, leading to DNA melting and assembly of the full replisome apparatus (1, 2). Key steps in MCM activation involve a series of phosphorylation-dependent events that promote the sequential association of Cdc45 and the GINS complex with the chromatin-associated MCM double hexamer. These recruitment events are regulated by the CDK and DDK kinases and require the additional accessory factors Sld3/7, Dpb11, and Sld2(3-6). The Cdc45•MCM(2-7)•GINS complex (CMG) forms the core of the eukaryotic replisome, and this 11-subunit assembly appears to be the functional helicase driving fork progression (3-6). The archaeal replication machinery resembles an ancestral form of its eukaryotic counterpart. Archaea possess a simple homohexameric MCM (5, 7). In addition, archaeal homologs of GINS and Cdc45 have been identified (8)(9)(10)(11)(12)(13)(14). In species of the genus Sulfolobus, we have previously demonstrated that the GINS complex interacts with the N-terminal domains of MCM (8). In Sulfolobus, GINS is a dimer of dimers: one subunit, Gins23, is related to the eukaryotic GINS components Psf2 and Psf3, and the second Sulfolobus subunit, Gins15, is related to the eukaryotic Sld5 and Psf1. These sequence relationships have been confirmed by structural studies of the Thermococcus kodakarensis GINS complex that have demonstrated the tetrameric assembly of archaeal (Gins15) 2 • (Gins23) 2 and validated the organizational similarity of the archaeal and eukaryotic GINS complexes (15). Interestingly, Sulfolobus GINS copurifies over the course of eight steps with a further polypeptide that we initially named RecJdbh, based on its observed homology with the presumptive DNA binding domain of the bacterial exonuclease, RecJ (8). Subsequent sequence analyses have revealed a relationship between RecJ and eukaryotic Cdc45, and this has been elegantly confirmed by recent structural studies of eukaryotic Cdc45 (9,11,16,17). We therefore propose renaming RecJdbh as Cdc45. As archaea lack orthologs of Sld2, Sld...

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“…One of them interacts with Hef (in this study) and the other interacts with GINS, an essential factor for DNA replication initiation and elongation in Eukarya and Archaea, which may be a component of the replicative helicase complex. The latter RecJ-like protein was reported as the GINS-associated nuclease (GAN), and may be involved in lagging strand processing (52). Furthermore, characterization of the GAN ortholog from P. furiosus revealed the 3Ј-5Ј exoribonuclease activity, which may function in proofreading for primer synthesis (53).…”

Section: Discussionmentioning

confidence: 99%

Multiple Interactions of the Intrinsically Disordered Region between the Helicase and Nuclease Domains of the Archaeal Hef Protein

Ishino

Yamagami

Kitamura

et al. 2014

Journal of Biological Chemistry

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Background: Hef/FANCM participates in interstrand cross-link DNA repair. Results: The predicted intrinsically disordered region (IDR) in Hef was experimentally verified. Proliferating cell nuclear antigen and a RecJ-like protein interact with the IDR individually, but not simultaneously. Conclusion:The IDR in Hef interacts with multiple proteins. Significance: Hef may function in DNA repair by association of its IDR with multiple partners.

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A novel DNA nuclease is stimulated by association with the GINS complex

Cited by 61 publications

References 34 publications

Archaeal DNA Polymerase D but Not DNA Polymerase B Is Required for Genome Replication in Thermococcus kodakarensis

Archaeal DNA Polymerase D but Not DNA Polymerase B Is Required for Genome Replication in Thermococcus kodakarensis

Archaeal orthologs of Cdc45 and GINS form a stable complex that stimulates the helicase activity of MCM

Multiple Interactions of the Intrinsically Disordered Region between the Helicase and Nuclease Domains of the Archaeal Hef Protein

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