Geminin Prevents Rereplication during Xenopus Development

To maintain a stable genome, it is essential that replication origins fire only once per cell cycle. The protein Geminin is thought to prevent a second round of DNA replication by inhibiting the essential replication factor Cdt1. Geminin also affects the development of several different organs by bi...

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Bibliographic Details
Published in:The Journal of biological chemistry 2007-02, Vol.282 (8), p.5514-5521
Main Authors: Kerns, Sarah L., Torke, Susanna J., Benjamin, Jacqueline M., McGarry, Thomas J.
Format: Article
Language:English
Subjects:
Animals
Blastula - cytology
Blastula - metabolism
Cell Cycle Proteins - genetics
Cell Cycle Proteins - metabolism
Chromatin Assembly and Disassembly - physiology
DNA Replication - physiology
DNA-Binding Proteins - genetics
DNA-Binding Proteins - metabolism
G2 Phase - physiology
Geminin
Gene Expression Regulation, Developmental - physiology
Mutation
Xenopus
Xenopus Proteins - genetics
Xenopus Proteins - metabolism
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Summary:To maintain a stable genome, it is essential that replication origins fire only once per cell cycle. The protein Geminin is thought to prevent a second round of DNA replication by inhibiting the essential replication factor Cdt1. Geminin also affects the development of several different organs by binding and inhibiting transcription factors and chromatin-remodeling proteins. It is not known if the defects in Geminin-deficient organisms are due to overreplication or to effects on cell differentiation. We previously reported that Geminin depletion in Xenopus causes early embryonic lethality due to a Chk1-dependent G2 cell cycle arrest just after the midblastula transition. Here we report that expressing a non-Geminin-binding Cdt1 mutant in Xenopus embryos exactly reproduces the phenotype of geminin depletion. Expressing the same mutant in replication extracts induces a partial second round of DNA replication within a single S phase. We conclude that Geminin is required to suppress a second round of DNA replication in vivo and that the phenotype of Geminin-depleted Xenopus embryos is caused by abnormal Cdt1 regulation. Expressing a nondegradable Cdt1 mutant in embryos also reproduces the Geminin-deficient phenotype. In cell extracts, the nondegradable mutant has no effect by itself but augments the amount of rereplication observed when Geminin is depleted. We conclude that Cdt1 is regulated both by Geminin binding and by degradation.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M609289200