Regulatory phosphorylation of the p34cdc2 protein kinase in vertebrates.
The p34cdc2 protein kinase is a conserved regulator of the eukaryotic cell cycle. Here we show that residues Thr14 and Tyr15 of mouse p34cdc2 become phosphorylated as mouse fibroblasts proceed through the cell cycle. We have mutated these residues and measured protein kinase activity of the p34cdc2 variants in a Xenopus egg extract. Phosphorylation of residues 14 and 15, which lie within the presumptive ATP-binding region of p34cdc2, normally restrains the protein kinase until it is specifically dephosphorylated and activated at the G2/M transition. Regulation by dephosphorylation of Tyr15 is conserved from fission yeast to mammals, while an extra level of regulation of mammalian p34cdc2 involves Thr14 dephosphorylation. In the absence of phosphorylation on these two residues, the kinase still requires cyclin B protein for its activation. Inhibition of DNA synthesis inhibits activation of wild-type p34cdc2 in the Xenopus system, but a mutant which cannot be phosphorylated at residues 14 and 15 escapes this inhibition, suggesting that these phosphorylation events form part of the pathway linking completion of DNA replication to initiation of mitosis.
Norbury
C
C
ICRF Cell Cycle Group, Department of Biochemistry, Oxford, UK.
Blow
J
J
Nurse
P
P
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001345:Autoradiography; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism*; D016213:Cyclins / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011233:Precipitin Tests; D006656:Protamine Kinase / Q000378:metabolism; D014981:Xenopus
D016213:Cyclins; D006656:Protamine Kinase; D016203:CDC2 Protein Kinase