Fan Heng-Yu, Tong Chao, Teng Chun-Bo, Lian Li, Li Shi-Wen, Yang Zeng-Ming, Chen Da-Yuan, Schatten Heide, Sun Qing-Yuan
State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, People's Republic of China.
Mol Reprod Dev. 2003 Jul;65(3):318-29. doi: 10.1002/mrd.10283.
Polo-like kinase 1 (Plk1) is a family of serine/threonine protein kinases that play important regulatory roles during mitotic cell cycle progression. In this study, Plk1 expression, subcellular localization, and possible functions during rat oocyte meiotic maturation, fertilization, and embryonic cleavages were studied by using RT-PCR, Western blot, confocal microscopy, drug-treatments, and antibody microinjection. Both the mRNA and protein of this kinase were detected in rat maturing oocytes and developing embryos. Confocal microscopy revealed that Plk1 distributed abundantly in the nucleus at the germinal vesicle (GV) stage, was associated with spindle poles during the formation of M-phase spindle, and was translocated to the spindle mid-zone at anaphase. In fertilized eggs, Plk1 was strongly stained in the cytoplasm between the apposing male and female pronuclei, from where microtubules radiated. Throughout cytokinesis, Plk1 was localized to the division plane, both during oocyte meiosis and embryonic mitosis. The specific subcellular distribution of Plk1 was distorted after disrupting the M-phase spindle, while additional aggregation dots could be induced in the cytoplasm by taxol, suggesting its intimate association with active microtubule assembly. Plk1 antibody microinjection delayed the meiotic resumption and blocked the emission of polar bodies. In conclusion, Plk1 may be a multifunctional kinase that plays pivotal regulatory roles in microtubule assembly during rat oocyte meiotic maturation, fertilization, and early embryonic mitosis.
Polo样激酶1(Plk1)是丝氨酸/苏氨酸蛋白激酶家族的一员,在有丝分裂细胞周期进程中发挥重要的调节作用。在本研究中,通过逆转录聚合酶链反应(RT-PCR)、蛋白质免疫印迹法(Western blot)、共聚焦显微镜技术、药物处理和抗体显微注射,研究了Plk1在大鼠卵母细胞减数分裂成熟、受精及胚胎卵裂过程中的表达、亚细胞定位及可能的功能。在大鼠成熟卵母细胞和发育中的胚胎中均检测到了该激酶的mRNA和蛋白质。共聚焦显微镜显示,在生发泡(GV)期,Plk1大量分布于细胞核中;在M期纺锤体形成过程中,Plk1与纺锤体两极相关联;在后期,Plk1转位至纺锤体中区。在受精卵中,Plk1在相对的雄原核和雌原核之间的细胞质中被强烈染色,微管从该区域发出。在整个胞质分裂过程中,无论是在卵母细胞减数分裂还是胚胎有丝分裂期间,Plk1均定位于分裂平面。破坏M期纺锤体后,Plk1的特定亚细胞分布发生扭曲,而紫杉醇可在细胞质中诱导额外的聚集点,提示其与活跃的微管组装密切相关。显微注射Plk1抗体可延迟减数分裂恢复并阻止极体排出。总之,Plk1可能是一种多功能激酶,在大鼠卵母细胞减数分裂成熟、受精及早期胚胎有丝分裂过程中的微管组装中发挥关键调节作用。