Norbury C, Blow J, Nurse P
ICRF Cell Cycle Group, Department of Biochemistry, Oxford, UK.
EMBO J. 1991 Nov;10(11):3321-9. doi: 10.1002/j.1460-2075.1991.tb04896.x.
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.
p34cdc2蛋白激酶是真核细胞周期中一种保守的调节因子。在此我们表明，随着小鼠成纤维细胞经历细胞周期，小鼠p34cdc2的苏氨酸14和酪氨酸15残基会发生磷酸化。我们已对这些残基进行突变，并在非洲爪蟾卵提取物中测定了p34cdc2变体的蛋白激酶活性。位于p34cdc2假定的ATP结合区域内的14和15位残基的磷酸化通常会抑制该蛋白激酶，直到它在G2/M期转换时被特异性去磷酸化并激活。从裂殖酵母到哺乳动物，酪氨酸15的去磷酸化调节都是保守的，而哺乳动物p34cdc2的另一层调节涉及苏氨酸14的去磷酸化。在这两个残基没有磷酸化的情况下，该激酶的激活仍需要细胞周期蛋白B。在非洲爪蟾系统中，DNA合成的抑制会抑制野生型p34cdc2的激活，但在14和15位残基不能被磷酸化的突变体则不受这种抑制，这表明这些磷酸化事件是将DNA复制的完成与有丝分裂的起始联系起来的信号通路的一部分。
