Skip to main content
SpringerLink
Log in

Activation of the MCM helicase from the thermophilic archaeon, Thermoplasma acidophilum by interactions with GINS and Cdc6-2

  • Special Issue: Original Paper
  • 10th International Congress on Extremophiles
  • Published:
Extremophiles Aims and scope Submit manuscript

Abstract

In DNA replication studies, the mechanism for regulation of the various steps from initiation to elongation is a crucial subject to understand cell cycle control. The eukaryotic minichromosome maintenance (MCM) protein complex is recruited to the replication origin by Cdc6 and Cdt1 to form the pre-replication complex, and participates in forming the CMG complex formation with Cdc45 and GINS to work as the active helicase. Intriguingly, Thermoplasma acidophilum, as well as many other archaea, has only one Gins protein homolog, contrary to the heterotetramer of the eukaryotic GINS made of four different proteins. The Gins51 protein reportedly forms a homotetramer (TaGINS) and physically interacts with TaMCM. In addition, TaCdc6-2, one of the two Cdc6/Orc1 homologs in T. acidophilum reportedly stimulates the ATPase and helicase activities of TaMCM in vitro. Here, we found a reaction condition, in which TaGINS stimulated the ATPase and helicase activities of TaMCM in a concentration dependent manner. Furthermore, the stimulation of the TaMCM helicase activity by TaGINS was enhanced by the addition of TaCdc6-2. A gel retardation assay revealed that TaMCM, TaGINS, and TaCdc6-2 form a complex on ssDNA. However, glutaraldehyde-crosslinking was necessary to detect the shifted band, indicating that the ternary complex of TaMCM–TaGINS–TaCdc6-2 is not stable in vitro. Immunoprecipitation experiment supported a weak interaction of these three proteins in vivo. Activation of the replicative helicase by a mechanism including a Cdc6-like protein suggests the divergent evolution after the division into Archaea and Eukarya.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Akita M, Adachi A, Takemura K, Yamagami T, Matsunaga F, Ishino Y (2010) Cdc6/Orc1 from Pyrococcus furiosus may act as the origin recognition protein and Mcm helicase recruiter. Genes Cells 15:537–552

    CAS  PubMed  Google Scholar 

  • Aparicio OM, Weinstein DM, Bell SP (1997) Components and dynamics of DNA replication complexes in S. cerevisiae: redistribution of MCM proteins and Cdc45p during S phase. Cell 91:59–69

    Article  CAS  PubMed  Google Scholar 

  • Chen YJ, Yu X, Kasiviswanathan R, Shin JH, Kelman Z, Egelman EH (2005) Structural polymorphism of Methanothermobacter thermautotrophicus MCM. J Mol Biol 346:389–394

    Article  CAS  PubMed  Google Scholar 

  • Costa A, van Duinen G, Medagli B, Chong J, Sakakibara N, Kelman Z, Nair SK, Patwardhan A, Onesti S (2008) Cryo-electron microscopy reveals a novel DNA-binding site on the MCM helicase. EMBO J 27:2250–2258

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Costa A, Ilves I, Tamberg N, Petojevic T, Nogales E, Botchan MR, Berger JM (2011) The structural basis for MCM2-7 helicase activation by GINS and Cdc45. Nat Struct Mol Biol 18:471–477

    Article  CAS  PubMed  Google Scholar 

  • Dueber EL, Corn JE, Bell SD, Berger JM (2007) Replication origin recognition and deformation by a heterodimeric archaeal Orc1 complex. Science 317:1210–1213

    Article  CAS  PubMed  Google Scholar 

  • Fu Y, Slaymaker IM, Wang J, Wang G, Chen XS (2013) The 1.8-Å crystal structure of the N-terminal domain of an Archaeal MCM as a right-handed filament. J Mol Biol 426:1512–1523

    Article  PubMed  Google Scholar 

  • Gambus A, Jones RC, Sanchez-Diaz A, Kanemaki M, van Deursen F, Edmondson RD, Labib K (2006) GINS maintains association of Cdc45 with MCM in replisome progression complexes at eukaryotic DNA replication forks. Nat Cell Biol 8:358–366

    Article  CAS  PubMed  Google Scholar 

  • Haugland GT, Shin JH, Birkeland NK, Kelman Z (2006) Stimulation of MCM helicase activity by a Cdc6 protein in the archaeon Thermoplasma acidophilum. Nucleic Acids Res 34:6337–6344

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Haugland GT, Innselset M, Madern D, Birkeland NK (2008a) Characterization of the Cdc6 Homologues from the Euryarchaeon Thermoplasma acidophilum. Open Biochem J 2:129–134

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Haugland GT, Sakakibara N, Pey AL, Rollor CR, Birkeland NK, Kelman Z (2008b) Thermoplasma acidophilum Cdc6 protein stimulates MCM helicase activity by regulating its ATPase activity. Nucleic Acids Res 36:5602–5609

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Haugland GT, Rollor CR, Birkeland NK, Kelman Z (2009) Biochemical characterization of the minichromosome maintenance protein from the archaeon Thermoplasma acidophilum. Extremophiles 13:81–88

    Article  CAS  PubMed  Google Scholar 

  • Ilves I, Petojevic T, Pesavento JJ, Botchan MR (2010) Activation of the MCM2-7 helicase by association with Cdc45 and GINS proteins. Mol Cell 37:247–258

    Article  CAS  PubMed  Google Scholar 

  • Ishimi Y (1997) A DNA helicase activity is associated with an MCM4, -6, and -7 protein complex. J Biol Chem 272:24508–24513

    Article  CAS  PubMed  Google Scholar 

  • Ishino Y, Ishino S (2012) Rapid progress of DNA replication studies in Archaea, the third domain of life. Sci China Ser-C Life Sci 55:1–18

    Article  Google Scholar 

  • Ishino S, Fujino S, Tomita H, Ogino H, Takao K, Daiyasu H, Kanai T, Atomi H, Ishino Y (2011) Biochemical and genetical analyses of the three mcm genes from the hyperthermophilic archaeon, Thermococcus kodakarensis. Genes Cells 16:1176–1189

    Article  CAS  PubMed  Google Scholar 

  • Kubota Y, Takase Y, Komori Y, Hashimoto Y, Arata T, Kamimura Y, Araki H, Takisawa H (2003) A novel ring-like complex of Xenopus proteins essential for the initiation of DNA replication. Genes Dev 17:1141–1152

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Leipe DD, Aravind L, Koonin EV (1999) Did DNA replication evolve twice independently? Nucleic Acids Res 27:3389–3401

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Li Z, Pan M, Santangelo TJ, Chemnitz W, Yuan W, Edwards JL, Hurwitz J, Reeve JN, Kelman Z (2011) A novel DNA nuclease is stimulated by association with the GINS complex. Nucleic Acids Res 39:6114–6123

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Makarova KS, Wolf YI, Mekhedov SL, Mirkin BG, Koonin EV (2005) Ancestral paralogs and pseudoparalogs and their role in the emergence of the eukaryotic cell. Nucleic Acids Res 33:4626–4638

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Makarova KS, Koonin EV, Kelman Z (2012) The CMG (CDC45/RecJ, MCM, GINS) complex is a conserved component of the DNA replication system in all archaea and eukaryotes. Biol Direct 7:7

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Marinsek N, Barry ER, Makarova KS, Dionne I, Koonin EV, Bell SD (2006) GINS, a central nexus in the archaeal DNA replication fork. EMBO Rep 7:539–545

    CAS  PubMed Central  PubMed  Google Scholar 

  • Masai H, Matsumoto S, You Z, Yoshizawa-Sugata N, Oda M (2010) Eukaryotic chromosome DNA replication: where, when, and how? Annu Rev Biochem 79:89–130

    Article  CAS  PubMed  Google Scholar 

  • Matsunaga F, Glatigny A, Mucchielli-Giorgi MH, Agier N, Delacroix H, Marisa L, Durosay P, Ishino Y, Aggerbeck L, Forterre P (2007) Genomewide and biochemical analyses of DNA-binding activity of Cdc6/Orc1 and Mcm proteins in Pyrococcus sp. Nucleic Acids Res 35:3214–3222

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Mott ML, Berger JM (2007) DNA replication initiation: mechanisms and regulation in bacteria. Nat Rev Microbiol 5:343–354

    Article  CAS  PubMed  Google Scholar 

  • Moyer SE, Lewis PW, Botchan MR (2006) Isolation of the Cdc45/Mcm2-7/GINS (CMG) complex, a candidate for the eukaryotic DNA replication fork helicase. Proc Natl Acad Sci USA 103:10236–10241

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Ogino H, Ishino S, Mayanagi K, Haugland GH, Birkeland N-K, Yamagishi A, Ishino Y (2011) The GINS complex from the thermoacidophilic archaeon, Thermoplasma acidophilum may function as a homotetramer in DNA replication. Extremophiles 15:529–539

    Article  CAS  PubMed  Google Scholar 

  • Pacek M, Tutter AV, Kubota Y, Takisawa H, Walter JC (2006) Localization of MCM2-7, Cdc45, and GINS to the site of DNA unwinding during eukaryotic DNA replication. Mol Cell 21:581–587

    Article  CAS  PubMed  Google Scholar 

  • Sakakibara N, Kelman LM, Kelman Z (2009) Unwinding the structure and function of the archaeal MCM helicase. Mol Microbiol 72:286–296

    Article  CAS  PubMed  Google Scholar 

  • Sanchez-Pulido L, Ponting CP (2011) Cdc45: the missing RecJ ortholog in eukaryotes? Bioinformatics 27:1885–1888

    Article  CAS  PubMed  Google Scholar 

  • Slaymaker IM, Fu Y, Toso DB, Ranatunga N, Brewster A, Forsburg SL, Zhou ZH, Chen XS (2013) Mini-chromosome maintenance complexes form a filament to remodel DNA structure and topology. Nucleic Acids Res 41:3446–3456

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Takayama Y, Kamimura Y, Okawa M, Muramatsu S, Sugino A, Araki H (2003) GINS, a novel multiprotein complex required for chromosomal DNA replication in budding yeast. Genes Dev 17:1153–1165

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Yasuda M, Oyaizu H, Yamagishi A, Oshima T (1995) Morphological variation of new Thermoplasma acidophilum isolates from Japanese hot springs. Appl Environ Microbiol 61:3482–3485

    CAS  PubMed Central  PubMed  Google Scholar 

  • Yoshimochi T, Fujikane R, Kawanami M, Matsunaga F, Ishino Y (2008) The GINS complex from Pyrococcus furiosus stimulates the MCM helicase activity. J Biol Chem 283:1601–1609

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan (Grant numbers 21113005, 23310152, and 26242075 to Y. I.). H. O. is supported by Grant-in-Aid for Japan Society for the Promotion of Science (JSPS) Fellows. This manuscript was written during N-K. Birkeland’s visit to Ishino’s laboratory by the Invitation Fellowship Programs for Research in Japan of JSPS.

Author information

Authors and Affiliations

  1. Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School of Bioresource and Bioenvironmental Sciences, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka, Fukuoka, 812-8581, Japan

    Hiromi Ogino, Sonoko Ishino & Yoshizumi Ishino

  2. Department of Biology, University of Bergen, Bergen, Norway

    Gyri Teien Haugland & Nils-Kåre Birkeland

  3. Medical Institute of Bioregulation, Kyushu University, Fukuoka, 812-8582, Japan

    Hiromi Ogino & Daisuke Kohda

Authors
  1. Hiromi Ogino
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sonoko Ishino
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Gyri Teien Haugland
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nils-Kåre Birkeland
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Daisuke Kohda
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Yoshizumi Ishino
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Yoshizumi Ishino.

Additional information

Communicated by M. W. W. Adams.

This article is part of a special issue based on the 10th International Congress on Extremophiles held in Saint Petersburg, Russia, September 7–11, 2014.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 402 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ogino, H., Ishino, S., Haugland, G.T. et al. Activation of the MCM helicase from the thermophilic archaeon, Thermoplasma acidophilum by interactions with GINS and Cdc6-2. Extremophiles 18, 915–924 (2014). https://doi.org/10.1007/s00792-014-0673-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00792-014-0673-6

Keywords

Search

Navigation