Cell cycle regulation of the replication licensing system: involvement of a Cdk-dependent inhibitor.
The replication licensing factor (RLF) is an essential initiation factor that is involved in preventing re-replication of chromosomal DNA in a single cell cycle. In Xenopus egg extracts, it can be separated into two components: RLF-M, a complex of MCM/P1 polypeptides, and RLF-B, which is currently unpurified. In this paper we investigate variations in RLF activity throughout the cell cycle. Total RLF activity is low in metaphase, due to a lack of RLF-B activity and the presence of an RLF inhibitor. RLF-B is rapidly activated on exit from metaphase, and then declines during interphase. The RLF inhibitor present in metaphase extracts is dependent on the activity of cyclin-dependent kinases (Cdks). Affinity depletion of Cdks from metaphase extracts removed the RLF inhibitor, while Cdc2/cyclin B directly inhibited RLF activity. In metaphase extracts treated with the protein kinase inhibitor 6-dimethylaminopurine (6-DMAP), both cyclin B and the RLF inhibitor were stabilized although the extracts morphologically entered interphase. These results are consistent with studies in other organisms that invoke a key role for Cdks in preventing re-replication of DNA in a single cell cycle.
Mahbubani
H M
HM
Imperial Cancer Research Fund, Clare Hall Laboratories, Potters Bar, Herts, United Kingdom.
Chong
J P
JP
Chevalier
S
S
Thömmes
P
P
Blow
J J
JJ
D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D018797:Cell Cycle Proteins / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D050759:Cyclin-Dependent Kinase Inhibitor p21; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000502:physiology*; D016213:Cyclins / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D004261:DNA Replication / Q000187:drug effects / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D014981:Xenopus
D018797:Cell Cycle Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004791:Enzyme Inhibitors; D011500:Protein Synthesis Inhibitors; C001020:N(6),N(6)-dimethyladenine; D003513:Cycloheximide; D018844:Cyclin-Dependent Kinases; D000225:Adenine