Elsevier

DNA Repair

Volume 72, December 2018, Pages 99-106
DNA Repair

Regulation of the initiation of DNA replication in human cells

https://doi.org/10.1016/j.dnarep.2018.09.003Get rights and content

Abstract

The origin of species would not have been possible without high fidelity DNA replication and complex genomes evolved with mechanisms that control the initiation of DNA replication at multiple origins on multiple chromosomes such that the genome is duplicated once and only once. The mechanisms that control the assembly and activation of the replicative helicase and the initiation of DNA replication in yeast and Xenopus egg extract systems have been identified and reviewed [1,2]. The goal of this review is to organize currently available data on the mechanisms that control the initiation of DNA replication in human cells.

Section snippets

Preface for a general audience

DNA replication is targeted by chemotherapeutic drugs as cancer cells generally proliferate faster than most normal cells, and many cancers have acquired mutations that inactivate mechanisms that ensure genome stability. A precise understanding of the mechanisms that initiate DNA replication will allow the rational design of clinical trials of new agents and combinations that target DNA replication. At this time our understanding of the mechanisms that initiate DNA replication is largely

Complexity and timing of the initiation of DNA replication

The human genome evolved with mechanisms that assemble and activate the replicative helicase to initiate DNA unwinding and replication at ∼50,000 origins (indirect estimations showed ∼22,000 in HeLa [3], ∼50,000 in HeLa Kyoto [4]). The MCM2-7 hexamer is loaded onto DNA to license origins in G1 phase and CDC7 (CDC7 functions in a heterodimer with DBF4 and is also known as DDK for DBF4-dependent kinase) and CDK2 kinase activities initiate the assembly of CDC45, MCM2-7 and GINS and activation of

Replication complex assembly

The assembly of the replisome in human cells and all eukaryotic systems begins with origin licensing in the G1 phase of the cell cycle (Fig. 1a).

MCM loading on chromatin in G1 phase human cells requires ORC (Origin Recognition Complex) [34,35], CDT1 [36] and CDC6 [37] (Table 1), and these proteins need to be tightly regulated to prevent re-loading of the replicative helicase and re-replication in S phase [38]. However, the mechanisms preventing re-replication are quite different between

The signaling required for the initiation of DNA replication

The kinases CDC7 [85,86] and CDK2 [72] play critical roles in the assembly and activation of pre-replication complexes in human cells (Fig. 1c, Table 2) and as such their regulation determines the initiation of DNA replication. CMG helicase activation in yeast and human cells requires MCM phosphorylation by DDK. DBF4-dependent kinase (DDK) (Fig. 2c, d) is a complex of CDC7 and either DBF4 or DBF4b (DRF1) [87]. DBF4 or DBF4b is required for maximum CDC7 kinase activity and substrate recognition.

Regulation of replication initiation after DNA damage

Key elements of the DNA damage response include cell cycle arrest and inhibition of the initiation of DNA replication (Fig. 3). The regulation of DNA replication after DNA damage is a very complex process. While mild replication stress activates the initiation of replication from dormant origins, higher levels of damage completely block origin firing. Current models for this phenomenon suggests that low levels of ATR/CHK1 activity block the activation of replication in the new “replication

Conclusions and future directions

The initiation of DNA replication in humans is an extremely complex process, and caution is necessary when attempting to translate findings from yeast and Xenopus extract systems into mammalian cells. Major gaps in knowledge about origin firing in human cells include the mechanism though which the GINS complex and DNA polymerase epsilon are recruited to the MCM helicase and the mechanism that regulates DDK after DNA damage. Also, while the requirement for CDK2 and CDC7 activities for certain

Conflicts of interest

The authors declare that there are no conflicts of interest.

Author contributions

TM and CJB wrote the paper.

Acknowledgement

This work was supported by the NIH Grant RO1 CA204173.

References (115)

  • Y. Jeon

    Human TopBP1 participates in cyclin E/CDK2 activation and preinitiation complex assembly during G1/S transition

    J. Biol. Chem.

    (2007)
  • Y. Gao

    Protein phosphatase 2A and Cdc7 kinase regulate the DNA unwinding element-binding protein in replication initiation

    J. Biol. Chem.

    (2014)
  • P. Perez-Arnaiz et al.

    An Mcm10 mutant defective in ssDNA binding shows defects in DNA replication initiation

    J. Mol. Biol

    (2016)
  • M. Izumi

    The Mcm2-7-interacting domain of human mini-chromosome maintenance 10 (Mcm10) protein is important for stable chromatin association and origin firing

    J. Biol. Chem.

    (2017)
  • V.P. Bermudez

    Studies on human DNA polymerase epsilon and GINS complex and their role in DNA replication

    J. Biol. Chem.

    (2011)
  • A.M. Sangoram

    Mammalian circadian autoregulatory loop: a timeless ortholog and mPer1 interact and negatively regulate CLOCK-BMAL1-induced transcription

    Neuron

    (1998)
  • N. Yoshizawa-Sugata et al.

    Human Tim/Timeless-interacting protein, Tipin, is required for efficient progression of S phase and DNA replication checkpoint

    J. Biol. Chem.

    (2007)
  • X. Xu

    TIMELESS suppresses the accumulation of aberrant CDC45.MCM2-7.GINS replicative helicase complexes on human chromatin

    J. Biol. Chem.

    (2016)
  • R. Kitamura

    Molecular mechanism of activation of human Cdc7 kinase: bipartite interaction with Dbf4/activator of S phase kinase (ASK) activation subunit stimulates ATP binding and substrate recognition

    J. Biol. Chem.

    (2011)
  • T. Silva

    Xenopus CDC7/DRF1 complex is required for the initiation of DNA replication

    J. Biol. Chem.

    (2006)
  • N. Yoshizawa-Sugata

    A second human Dbf4/ASK-related protein, Drf1/ASKL1, is required for efficient progression of S and M phases

    J. Biol. Chem.

    (2005)
  • A. Ballabeni

    Human CDT1 associates with CDC7 and recruits CDC45 to chromatin during S phase

    J. Biol. Chem.

    (2009)
  • K.A. Merrick

    Distinct activation pathways confer cyclin-binding specificity on Cdk1 and Cdk2 in human cells

    Mol. Cell.

    (2008)
  • J.P. Welburn

    How tyrosine 15 phosphorylation inhibits the activity of cyclin-dependent kinase 2-cyclin A

    J. Biol. Chem.

    (2007)
  • C. Berthet

    Cdk2 knockout mice are viable

    Curr. Biol.

    (2003)
  • K.A. Merrick

    Switching Cdk2 on or off with small molecules to reveal requirements in human cell proliferation

    Mol. Cell.

    (2011)
  • S.P. Bell et al.

    Chromosome duplication in Saccharomyces cerevisiae

    Genetics

    (2016)
  • J.J. Blow et al.

    Xenopus cell-free extracts and their contribution to the study of DNA replication and other complex biological processes

    Int. J. Dev. Biol.

    (2016)
  • D.A. Jackson et al.

    Replicon clusters are stable units of chromosome structure: evidence that nuclear organization contributes to the efficient activation and propagation of S phase in human cells

    J. Cell. Biol.

    (1998)
  • V.O. Chagin

    4D visualization of replication foci in mammalian cells corresponding to individual replicons

    Nat. Commun.

    (2016)
  • M. O’Donnell et al.

    Principles and concepts of DNA replication in bacteria, archaea, and eukarya

    Cold Spring Harb. Perspect. Biol

    (2013)
  • J. Ferreira

    Spatial organization of large-scale chromatin domains in the nucleus: a magnified view of single chromosome territories

    J Cell. Biol.

    (1997)
  • J. Gerhardt

    Identification of new human origins of DNA replication by an origin-trapping assay

    Mol. Cell. Biol

    (2006)
  • N. Petryk

    Replication landscape of the human genome

    Nat. Commun.

    (2016)
  • F. Picard

    The spatiotemporal program of DNA replication is associated with specific combinations of chromatin marks in human cells

    PLoS Genet.

    (2014)
  • H.M. Mahbubani

    Cell cycle regulation of the replication licensing system: involvement of a Cdk-dependent inhibitor

    J. Cell. Biol

    (1997)
  • S. Donovan

    Cdc6p-dependent loading of Mcm proteins onto pre-replicative chromatin in budding yeast

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

    (1997)
  • R. Burkhart

    Interactions of human nuclear proteins P1Mcm3 and P1Cdc46

    Eur. J. Biochem.

    (1995)
  • A. Ibarra et al.

    Excess MCM proteins protect human cells from replicative stress by licensing backup origins of replication

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

    (2008)
  • X.Q. Ge et al.

    Dormant origins licensed by excess Mcm2-7 are required for human cells to survive replicative stress

    Genes Dev.

    (2007)
  • M. Macheret et al.

    Intragenic origins due to short G1 phases underlie oncogene-induced DNA replication stress

    Nature

    (2018)
  • D. Lob

    3D replicon distributions arise from stochastic initiation and domino-like DNA replication progression

    Nat. Commun.

    (2016)
  • Y. Gindin

    A chromatin structure-based model accurately predicts DNA replication timing in human cells

    Mol. Syst. Biol.

    (2014)
  • B. Miotto et al.

    Selectivity of ORC binding sites and the relation to replication timing, fragile sites, and deletions in cancers

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

    (2016)
  • O. Hyrien

    How MCM loading and spreading specify eukaryotic DNA replication initiation sites

    F1000Res

    (2016)
  • S.C. Yang et al.

    Modeling genome-wide replication kinetics reveals a mechanism for regulation of replication timing

    Mol. Syst. Biol.

    (2010)
  • S.P. Das

    Replication timing is regulated by the number of MCMs loaded at origins

    Genome Res.

    (2015)
  • S. Yamazaki

    Rif1 regulates the replication timing domains on the human genome

    EMBO J.

    (2012)
  • J.R. Dixon

    Topological domains in mammalian genomes identified by analysis of chromatin interactions

    Nature

    (2012)
  • B.D. Pope

    Topologically associating domains are stable units of replication-timing regulation

    Nature

    (2014)
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