Temporal regulation of cdc2 mitotic kinase activity and cyclin degradation in cell-free extracts of Xenopus eggs.
In cleaving Xenopus eggs, the cell division cycle is abbreviated to a rapid succession of S and M phases. During mitosis a number of proteins show increased phosphorylation due to the activation of a histone H1 kinase, the homologue of the cdc2+ gene product of the yeast Schizosaccharomyces pombe. We have studied the regulation of the activity of this enzyme in cell-free extracts of Xenopus eggs. In extracts of activated eggs incubated at 22 degrees C, histone H1 kinase activity shows two peaks of activation and disappearance. Activation occurs in two stages. The first stage requires protein synthesis, whereas the second does not. The second stage of activation involves post-translational activation of the kinase. Kinase activity rises to a peak and then abruptly disappears. Added sea urchin cyclin is degraded at the time of disappearance of kinase activity. The oscillation in kinase activity is then repeated, usually with lower amplitude. Post-translational activation of the kinase requires a membrane-containing particulate cellular component, whose role has yet to be defined. The kinase can still be activated in the presence of EDTA or in the presence of the ATP analogue, 6-dimethylaminopurine, which implies that phosphorylation of the kinase complex is not required for activation. Under these conditions, however, the kinase activity does not show its normal sudden disappearance, and added cyclin is perfectly stable. These observations are consistent with the idea that post-translational activation of the kinase involves protein phosphatase activity, whereas switching off the kinase requires an ATP-Mg2(+)-dependent reaction, perhaps due to protein phosphorylation
Felix
M A
MA
European Molecular Biology Laboratory, Heidelberg, FRG.
Pines
J
J
Hunt
T
T
Karsenti
E
E
D000225:Adenine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D016203:CDC2 Protein Kinase; D016213:Cyclins; D004492:Edetic Acid / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D005260:Female; D007447:Invertebrate Hormones / Q000378:metabolism*; D008938:Mitosis; D009687:Nuclear Proteins / Q000378:metabolism*; D010063:Ovum; D010750:Phosphoproteins / Q000378:metabolism*; D006656:Protamine Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D013997:Time Factors; D014982:Xenopus laevis
D016213:Cyclins; D007447:Invertebrate Hormones; D009687:Nuclear Proteins; D010750:Phosphoproteins; C001020:N(6),N(6)-dimethyladenine; D004492:Edetic Acid; D011494:Protein Kinases; D006656:Protamine Kinase; D016203:CDC2 Protein Kinase; D000225:Adenine