Erikson E, Maller J L
Department of Pharmacology, University of Colorado School of Medicine, Denver 80262.
J Biol Chem. 1989 Aug 15;264(23):13711-7.
Previous studies have shown that increased ribosomal protein S6 kinase activity in unfertilized Xenopus eggs can be resolved by DEAE-Sephacel chromatography into two peaks, designated S6 kinase I and S6 kinase II. We show here that antibody against bacterially expressed S6 kinase II cross-reacts with S6 kinase I. Both S6 kinases undergo marked phosphorylation when they are activated during oocyte maturation, and both become deactivated and dephosphorylated upon activation of eggs. Immunoblotting of extracts of oocytes reveals that all S6 kinase molecules undergo a decrease and increase in electrophoretic mobility upon activation and deactivation, respectively. The increase in electrophoretic mobility can be produced in vitro by incubation of activated S6 kinase with purified phosphatases. Phosphoamino acid analysis of S6 kinase II labeled in vivo during maturation reveals both phosphoserine and phosphothreonine, and phosphopeptide maps suggest that several kinases may phosphorylate and activate S6 kinase II in vivo. These results demonstrate that, during oocyte maturation and early development, S6 kinase activation and deactivation are regulated by phosphorylation and dephosphorylation, suggesting a probable mechanism for S6 kinase regulation in other mitogenically stimulated cells.
先前的研究表明，未受精的非洲爪蟾卵中核糖体蛋白S6激酶活性的增加可通过DEAE-葡聚糖凝胶层析分离为两个峰，分别称为S6激酶I和S6激酶II。我们在此表明，针对细菌表达的S6激酶II的抗体与S6激酶I发生交叉反应。在卵母细胞成熟过程中激活时，两种S6激酶都会发生明显的磷酸化，而在卵子激活时都会失活并去磷酸化。对卵母细胞提取物进行免疫印迹分析表明，所有S6激酶分子在激活和失活时电泳迁移率分别下降和增加。通过将激活的S6激酶与纯化的磷酸酶一起孵育，可以在体外产生电泳迁移率的增加。对成熟过程中体内标记的S6激酶II进行磷酸氨基酸分析，发现了磷酸丝氨酸和磷酸苏氨酸，磷酸肽图谱表明几种激酶可能在体内磷酸化并激活S6激酶II。这些结果表明，在卵母细胞成熟和早期发育过程中，S6激酶的激活和失活受磷酸化和去磷酸化调节，这提示了其他有丝分裂原刺激细胞中S6激酶调节的可能机制。
