Walter S A, Guadagno T M, Ferrell J E
Department of Molecular Pharmacology, Stanford University School of Medicine, California 94305-5332, USA.
Mol Biol Cell. 1997 Nov;8(11):2157-69. doi: 10.1091/mbc.8.11.2157.
Previous work has established that activation of Mos, Mek, and p42 mitogen-activated protein (MAP) kinase can trigger release from G2-phase arrest in Xenopus oocytes and oocyte extracts and can cause Xenopus embryos and extracts to arrest in mitosis. Herein we have found that activation of the MAP kinase cascade can also bring about an interphase arrest in cycling extracts. Activation of the cascade early in the cycle was found to bring about the interphase arrest, which was characterized by an intact nuclear envelope, partially condensed chromatin, and interphase levels of H1 kinase activity, whereas activation of the cascade just before mitosis brought about the mitotic arrest, with a dissolved nuclear envelope, condensed chromatin, and high levels of H1 kinase activity. Early MAP kinase activation did not interfere significantly with DNA replication, cyclin synthesis, or association of cyclins with Cdc2, but it did prevent hyperphosphorylation of Cdc25 and Wee1 and activation of Cdc2/cyclin complexes. Thus, the extracts were arrested in a G2-like state, unable to activate Cdc2/cyclin complexes. The MAP kinase-induced G2 arrest appeared not to be related to the DNA replication checkpoint and not to be mediated through inhibition of Cdk2/cyclin E; evidently a novel mechanism underlies this arrest. Finally, we found that by delaying the inactivation of MAP kinase during release of a cytostatic factor-arrested extract from its arrest state, we could delay the subsequent entry into mitosis. This finding suggests that it is the persistence of activated MAP kinase after fertilization that allows the occurrence of a G2-phase during the first mitotic cell cycle.
先前的研究已经证实，Mos、Mek和p42丝裂原活化蛋白（MAP）激酶的激活可触发非洲爪蟾卵母细胞和卵母细胞提取物从G2期阻滞中释放，并可导致非洲爪蟾胚胎和提取物在有丝分裂期发生阻滞。在此，我们发现MAP激酶级联的激活也可导致循环提取物在间期发生阻滞。研究发现，在细胞周期早期激活该级联会导致间期阻滞，其特征为核膜完整、染色质部分凝聚以及H1激酶活性处于间期水平，而在有丝分裂前激活该级联则会导致有丝分裂期阻滞，核膜溶解、染色质凝聚且H1激酶活性水平较高。早期MAP激酶激活对DNA复制、细胞周期蛋白合成或细胞周期蛋白与Cdc2的结合没有显著干扰，但它确实阻止了Cdc25和Wee1的过度磷酸化以及Cdc2/细胞周期蛋白复合物的激活。因此，提取物停滞在类似G2的状态，无法激活Cdc2/细胞周期蛋白复合物。MAP激酶诱导的G2期阻滞似乎与DNA复制检查点无关，也不是通过抑制Cdk2/细胞周期蛋白E介导的；显然，这种阻滞有一个新的机制。最后，我们发现，通过在从细胞静止因子阻滞状态释放提取物的过程中延迟MAP激酶的失活，我们可以延迟随后进入有丝分裂期。这一发现表明，受精后活化MAP激酶的持续存在使得在第一个有丝分裂细胞周期中出现G2期。
