Center for Reproductive Medicine, Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, China.
Cell Cycle. 2011 Jun 1;10(11):1861-70. doi: 10.4161/cc.10.11.15797.
GM130, a cis-Golgi protein, plays key roles in various mitotic events, but its function in mammalian oocyte meiosis remains unknown. In this study, we found that GM130 was localized to the spindle poles at both metaphase I and metaphase II stages and associated with the midbody at telophase I stage. The association of GM130 with spindle poles was further confirmed by its colocalization with the centrosome-associated proteins, MEK1/2. By nocodazole treatment, we clarified that GM130 localization was consistently dependent on spindle assembly. Then we investigated the possible function of GM130 by specific morpholino microinjection. This treatment caused abnormal spindle formation, and decreased first polar body extrusion. Our results showed that knockdown of GM130 impaired the localization of MTOCs proteins γ-tubulin and Plk1. Using live cell imaging we observed that depletion of GM130 affected spindle migration and resulted in elongated spindle and large polar body extrusion. We further found that depletion of GM130 blocked p-MEK1/2 accumulation at the spindle poles. And, it was shown that GM130 detached from the spindle poles in oocytes treated with MEK specific inhibitor U0126. Taken together, our results suggested that GM130 regulates microtubule organization and might cooperate with the MAPK pathway to play roles in spindle organization, migration and asymmetric division during mouse oocyte maturation.
GM130 是一种顺式高尔基蛋白,在各种有丝分裂事件中发挥关键作用,但在哺乳动物卵母细胞减数分裂中的功能尚不清楚。在本研究中,我们发现 GM130 定位于中期 I 和中期 II 的纺锤体两极,并与末期 I 的中体相关联。GM130 与纺锤体两极的关联通过其与中心体相关蛋白 MEK1/2 的共定位进一步得到证实。通过用诺考达唑处理,我们澄清 GM130 的定位始终依赖于纺锤体组装。然后,我们通过特异性的 morpholino 微注射研究 GM130 的可能功能。这种处理导致异常的纺锤体形成,并减少第一极体的挤出。我们的结果表明,GM130 的敲低破坏了 MTOC 蛋白 γ-微管蛋白和 Plk1 的定位。通过活细胞成像观察到,GM130 的耗竭影响纺锤体的迁移,导致纺锤体拉长和大极体挤出。我们进一步发现,GM130 的耗竭阻止了 MEK 特异性抑制剂 U0126 处理的卵母细胞中 p-MEK1/2 在纺锤体两极的积累。此外,还表明 GM130 在 U0126 处理的卵母细胞中从纺锤体两极脱离。综上所述,我们的结果表明 GM130 调节微管组织,并可能与 MAPK 途径合作,在小鼠卵母细胞成熟过程中发挥纺锤体组织、迁移和不对称分裂的作用。