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Published online 19 April 2004. doi:10.1083/jcb.200311044
The Rockefeller University Press, 0021-9525 $8.00
JCB, Volume 165, Number 2, 181-190
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Article

 The role of Cdc6 in ensuring complete genome licensing and S phase checkpoint activation

Maren Oehlmann, Alan J. Score, and J. Julian Blow

Wellcome Trust Biocentre, University of Dundee, Dundee, UK

Address correspondence to J. Julian Blow, Wellcome Trust Biocentre, University of Dundee, Dow Street, Dundee DD1 5EH, UK. Tel.: 44-1-382-345-797. Fax: 44-1-382-348-072. email: j.j.blow{at}dundee.ac.uk

Before S phase, cells license replication origins for initiation by loading them with Mcm2-7 heterohexamers. This process is dependent on Cdc6, which is recruited to unlicensed origins. Using Xenopus egg extracts we show that although each origin can load many Mcm2-7 hexamers, the affinity of Cdc6 for each origins drops once it has been licensed by loading the first hexamers. This encourages the distribution of at least one Mcm2-7 hexamer to each origin, and thereby helps to ensure that all origins are licensed. Although Cdc6 is not essential for DNA replication once licensing is complete, Cdc6 regains a high affinity for origins once replication forks are initiated and Mcm2-7 has been displaced from the origin DNA. We show that the presence of Cdc6 during S phase is essential for the checkpoint kinase Chk1 to become activated in response to replication inhibition. These results show that Cdc6 plays multiple roles in ensuring precise chromosome duplication.

Key Words: DNA replication; Cdc6; replication licensing; Xenopus; Chk1

Abbreviations used in this paper: NIB, nuclear isolation buffer; ORC, origin recognition complex; pre-RC, pre-replicative complex.


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