Wang Yanzhuang, Seemann Joachim, Pypaert Marc, Shorter James, Warren Graham
Department of Cell Biology, Ludwig Institute for Cancer Research, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520, USA.
EMBO J. 2003 Jul 1;22(13):3279-90. doi: 10.1093/emboj/cdg317.
Cell-free assays that mimic the disassembly and reassembly cycle of the Golgi apparatus during mitosis implicated GRASP65 as a mitotically regulated stacking factor. We now present evidence that GRASP65 is directly involved in stacking Golgi cisternae. GRASP65 is the major phosphorylation target in rat liver Golgi membranes of two mitotic kinases, cdc2-cyclin B and polo-like kinases, which alone will unstack Golgi membranes, generating single cisternae. Mitotic cells microinjected with antibodies to GRASP65 fail to form proper Golgi stacks after cell division. Beads coated with GRASP65 homodimers form extensive aggregates consistent with the formation of trans oligomers. These can be disaggregated using purified cdc2-cyclin B1 and polo-like kinases, and re-aggregated after dephosphorylation of GRASP65. Together, these data demonstrate that GRASP65 has the properties required to bind surfaces together in a mitotically regulated manner.
在有丝分裂期间模拟高尔基体拆卸和重新组装循环的无细胞分析表明，GRASP65是一种受有丝分裂调控的堆叠因子。我们现在提供证据表明，GRASP65直接参与高尔基体潴泡的堆叠。GRASP65是大鼠肝脏高尔基体膜中两种有丝分裂激酶（cdc2 - 细胞周期蛋白B和polo样激酶）的主要磷酸化靶点，这两种激酶单独作用会使高尔基体膜解堆叠，产生单个潴泡。微注射GRASP65抗体的有丝分裂细胞在细胞分裂后无法形成正常的高尔基体堆叠。涂有GRASP65同型二聚体的珠子形成大量聚集体，这与反式寡聚体的形成一致。使用纯化的cdc2 - 细胞周期蛋白B1和polo样激酶可以使这些聚集体解聚，并在GRASP65去磷酸化后重新聚集。这些数据共同表明，GRASP65具有以有丝分裂调控方式将表面结合在一起所需的特性。
