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Vol. 13, Issue 12, 4388-4400, December 2002

Identification of the Nuclear Localization Signal in Xenopus Cyclin E and Analysis of Its Role in Replication and Mitosis

Jonathan D. Moore,^*¶ Sally Kornbluth, and Tim Hunt^*

 ^*Cancer Research UK London Research Institute, Clare Hall Laboratories, South Mimms, Herts, EN6 3LD, United Kingdom, and  Department of Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina, 27710-3686

Cyclin-dependent kinase (Cdk)2/cyclin E is imported into nuclei assembled in Xenopus egg extracts by a pathway that requires importin- and -. Here, we identify a basic nuclear localization sequence (NLS) in the N-terminus of Xenopus cyclin E. Mutation of the NLS eliminated nuclear accumulation of both cyclin E and Cdk2, and such versions of cyclin E were unable to trigger DNA replication. Addition of a heterologous NLS from SV40 large T antigen restored both nuclear targeting of Cdk2/cyclin E and DNA replication. We present evidence indicating that Cdk2/cyclin E complexes must become highly concentrated within nuclei to support replication and find that cyclin A can trigger replication at much lower intranuclear concentrations. We confirmed that depletion of endogenous cyclin E increases the concentration of cyclin B necessary to promote entry into mitosis. In contrast to its inability to promote DNA replication, cyclin E lacking its NLS was able to cooperate with cyclin B in promoting mitotic entry.

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Corresponding author. E-mail address: tim.hunt{at}cancer.org.uk.

^¶ Present address: Ribotargets Ltd., Granta Park, Cambridge, CB1 6GB, England.

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Molecular Biology of the Cell
Vol. 13, 4388-4400, December 2002
Copyright © 2002 by The American Society for Cell Biology

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