Sun B, Murray N R, Fields A P
Sealy Center for Oncology and Hematology, University of Texas Medical Branch, Galveston, Texas 77555-1048, USA.
J Biol Chem. 1997 Oct 17;272(42):26313-7. doi: 10.1074/jbc.272.42.26313.
Protein kinase C (PKC) is activated at the nucleus during the G2 phase of cell cycle, where it is required for mitosis. However, the mechanisms controlling cell cycle-dependent activation of nuclear PKC are not known. We now report that nuclear levels of the major physiologic PKC activator diacylglycerol (DAG) fluctuate during cell cycle. Specifically, nuclear DAG levels in G2/M phase cells are 2. 5-3-fold higher than in G1 phase cells. In synchronized cells, nuclear DAG levels rise to a peak coincident with the G2/M phase transition and return to basal levels in G1 phase cells. This increase in DAG level is sufficient to stimulate betaII PKC-mediated phosphorylation of its mitotic nuclear envelope substrate lamin B in vitro. Isolated nuclei from G2 phase cells contain an active phospholipase activity capable of generating DAG in vitro. Nuclear phospholipase activity is inhibited by the selective phosphatidylinositol-specific phospholipase C (PI-PLC) inhibitor 1-O-octadeyl-2-O-methyl-sn-glycero-3-phosphocholine and neomycin sulfate, but not by the phosphatidylcholine-PLC selective inhibitor D609 or inhibitors of phospholipase D-mediated DAG generation. Treatment of synchronized cells with 1-O-octadeyl-2-O-methyl-sn-glycero-3-phosphocholine leads to decreased nuclear PI-PLC activity and cell cycle blockade in the G2 phase, suggesting a role for nuclear PI-PLC in the G2/M phase transition. Our data are consistent with the hypothesis that nuclear PI-PLC generates DAG to activate nuclear betaII PKC, whose activity is required for mitosis.
蛋白激酶C（PKC）在细胞周期的G2期于细胞核内被激活，而有丝分裂需要这种激活作用。然而，控制细胞核PKC细胞周期依赖性激活的机制尚不清楚。我们现在报告，主要生理性PKC激活剂二酰基甘油（DAG）的细胞核水平在细胞周期中会发生波动。具体而言，G2/M期细胞中的细胞核DAG水平比G1期细胞高2.5至3倍。在同步化细胞中，细胞核DAG水平上升至峰值，与G2/M期转换同时出现，并在G1期细胞中恢复至基础水平。DAG水平的这种升高足以在体外刺激βII PKC介导的有丝分裂核膜底物核纤层蛋白B的磷酸化。来自G2期细胞的分离细胞核含有一种活性磷脂酶活性，能够在体外生成DAG。细胞核磷脂酶活性受到选择性磷脂酰肌醇特异性磷脂酶C（PI-PLC）抑制剂1-O-十八烷基-2-O-甲基-sn-甘油-3-磷酸胆碱和硫酸新霉素的抑制，但不受磷脂酰胆碱-PLC选择性抑制剂D609或磷脂酶D介导的DAG生成抑制剂的抑制。用1-O-十八烷基-2-O-甲基-sn-甘油-3-磷酸胆碱处理同步化细胞会导致细胞核PI-PLC活性降低以及细胞周期在G2期阻滞，这表明细胞核PI-PLC在G2/M期转换中发挥作用。我们的数据与以下假设一致，即细胞核PI-PLC生成DAG以激活细胞核βII PKC，而有丝分裂需要其活性。
