Roles of the Golgi apparatus in cell growth regulation

GRASP65在S277被cdc2/Cyclin B磷酸化，可能参与有丝分裂进入的调节。通过两种方法分析GRASP65S277磷酸化在细胞周期调控中的作用。(1)将不能定位于高尔基体并聚集在细胞质中的重组GRASP65G2a突变体注入细胞内，可抑制早期组蛋白H3的磷酸化，但不能抑制中心体的分离。这一结果提示，组蛋白H3的磷酸化和中心体的分离在有丝分裂开始时是独立调节的，而GRASP65的S277磷酸化可能在这些事件的调节中起一定作用。(2)生化分析表明，PLK-1与GRASP65结合是通过有丝分裂e的磷酸化，这依赖于S277的磷酸化。这种结合是由Plk1的Polo-box结构域介导的。这一结果表明，GRASP65在S277处被磷酸化，招募Plk1，并在有丝分裂开始时作为信号支架发挥作用。

GRASP65 is phosphorylated at S277 by cdc2/cyclin B and may participate in the regulation of mitotic entry. The role of GRASP65 S277 phosphorylation in cell cycle regulation was analyzed by two approaches. (1) Injection of recombinant GRASP65 G2A mutant, which cannot localize to the Golgi apparatus and accumulate in the cytoplasm, into the cells inhibited the phosphorylation of histone H3 in early prophase but not the centrosome separation. This result suggested that phosphorylation of histone H3 and centrosome separation are independently regulated at the onset of mitosis and S277 phosphorylation of GRASP65 may have some role in the regulation of those events. (2) Biochemical analyses revealed that Plk 1 binds to GRASP65 phosphorylated by mitotic e and this was dependent on the phosphorylation of S277. The binding is mediated by polo-box domain of Plk1. This result suggested that GRASP65 phosphorylated at S277 recruit Plk1 and function as a signaling scaffold at the onset of mitosis.