Identification of the nuclear localization signal in Xenopus cyclin E and analysis of its role in replication and mitosis.
Cyclin-dependent kinase (Cdk)2/cyclin E is imported into nuclei assembled in Xenopus egg extracts by a pathway that requires importin-alpha and -beta. 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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Related Subject Headings
- Xenopus Proteins
- Xenopus
- Sequence Homology, Amino Acid
- Protein Structure, Tertiary
- Protein Serine-Threonine Kinases
- Plasmids
- Nuclear Localization Signals
- Mutation
- Molecular Sequence Data
- Mitosis
Citation
Published In
DOI
EISSN
ISSN
Publication Date
Volume
Issue
Start / End Page
Related Subject Headings
- Xenopus Proteins
- Xenopus
- Sequence Homology, Amino Acid
- Protein Structure, Tertiary
- Protein Serine-Threonine Kinases
- Plasmids
- Nuclear Localization Signals
- Mutation
- Molecular Sequence Data
- Mitosis