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  The ORC/Cdc6/MCM2-7 complex facilitates MCM2-7 dimerization during prereplicative complex formation

Cecile, E., Fernandez-Cid, A., Riera, A., Zech, J., Clarke, P., Herrera, M. C., et al. (2014). The ORC/Cdc6/MCM2-7 complex facilitates MCM2-7 dimerization during prereplicative complex formation. Nucleic Acids Research (London), 42(4), 2257-2269. doi:10.1093/nar/gkt1148.

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Cecile , Evrin1, Author
Fernandez-Cid, Alejandra 1, Author
Riera, Alberto1, Author
Zech, Jürgen1, Author
Clarke, Pippa1, Author
Herrera, M. Carmen 1, Author
Tognetti, Silvia 1, Author
Lurz, Rudi2, Author              
Speck, Christian 1, Author
Affiliations:
1DNA Replication Group, MRC Clinical Sciences Centre, Imperial College, Du Cane Road, London W12 0NN, UK, ou_persistent22              
2Imaging/Electron Microscopy (Head: Thorsten Mielke), Scientific Service (Head: Christoph Krukenkamp), Max Planck Institute for Molecular Genetics, Max Planck Society, ou_1479668              

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 Abstract: The replicative mini-chromosome-maintenance 2-7 (MCM2-7) helicase is loaded in Saccharomyces cerevisiae and other eukaryotes as a head-to-head double-hexamer around origin DNA. At first, ORC/Cdc6 recruits with the help of Cdt1 a single MCM2-7 hexamer to form an 'initial' ORC/Cdc6/Cdt1/MCM2-7 complex. Then, on ATP hydrolysis and Cdt1 release, the 'initial' complex is transformed into an ORC/Cdc6/MCM2-7 (OCM) complex. However, it remains unclear how the OCM is subsequently converted into a MCM2-7 double-hexamer. Through analysis of MCM2-7 hexamer-interface mutants we discovered a complex competent for MCM2-7 dimerization. We demonstrate that these MCM2-7 mutants arrest during prereplicative complex (pre-RC) assembly after OCM formation, but before MCM2-7 double-hexamer assembly. Remarkably, only the OCM complex, but not the 'initial' ORC/Cdc6/Cdt1/MCM2-7 complex, is competent for MCM2-7 dimerization. The MCM2-7 dimer, in contrast to the MCM2-7 double-hexamer, interacts with ORC/Cdc6 and is salt-sensitive, classifying the arrested complex as a helicase-loading intermediate. Accordingly, we found that overexpression of the mutants cause cell-cycle arrest and dominant lethality. Our work identifies the OCM complex as competent for MCM2-7 dimerization, reveals MCM2-7 dimerization as a limiting step during pre-RC formation and defines critical mechanisms that explain how origins are licensed.

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Language(s): eng - English
 Dates: 2013-10-252013-11-142014-02-01
 Publication Status: Published in print
 Pages: 13
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 Table of Contents: -
 Rev. Type: -
 Identifiers: DOI: 10.1093/nar/gkt1148
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Title: Nucleic Acids Research (London)
  Other : Nucleic Acids Res
Source Genre: Journal
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Publ. Info: Oxford : Oxford University Press
Pages: - Volume / Issue: 42 (4) Sequence Number: - Start / End Page: 2257 - 2269 Identifier: ISSN: 0305-1048
CoNE: https://pure.mpg.de/cone/journals/resource/110992357379342