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RanGTP and importin β regulate meiosis I spindle assembly and function in mouse oocytes.
The EMBO Journal ( IF 9.4 ) Pub Date : 2019-10-16 , DOI: 10.15252/embj.2019101689
David Drutovic 1 , Xing Duan 2, 3 , Rong Li 2, 3 , Petr Kalab 2 , Petr Solc 1
Affiliation  

Homologous chromosome segregation during meiosis I (MI) in mammalian oocytes is carried out by the acentrosomal MI spindles. Whereas studies in human oocytes identified Ran GTPase as a crucial regulator of the MI spindle function, experiments in mouse oocytes questioned the generality of this notion. Here, we use live-cell imaging with fluorescent probes and Förster resonance energy transfer (FRET) biosensors to monitor the changes in Ran and importin β signaling induced by perturbations of Ran in mouse oocytes while examining the MI spindle dynamics. We show that unlike RanT24N employed in previous studies, a RanT24N, T42A double mutant inhibits RanGEF without perturbing cargo binding to importin β and disrupts MI spindle function in chromosome segregation. Roles of Ran and importin β in the coalescence of microtubule organizing centers (MTOCs) and MI spindle assembly are further supported by the use of the chemical inhibitor importazole, whose effects are partially rescued by the GTP hydrolysis-resistant RanQ69L mutant. These results indicate that RanGTP is essential for MI spindle assembly and function both in humans and mice.

中文翻译:


RanGTP 和 importin β 调节小鼠卵母细胞减数分裂 I 纺锤体的组装和功能。



哺乳动物卵母细胞减数分裂 I (MI) 期间的同源染色体分离是由心体 MI 纺锤体进行的。尽管对人类卵母细胞的研究发现 Ran GTPase 是 MI 纺锤体功能的关键调节因子,但对小鼠卵母细胞的实验却对这一概念的普遍性提出了质疑。在这里,我们使用荧光探针和 Förster 共振能量转移 (FRET) 生物传感器的活细胞成像来监测小鼠卵母细胞中 Ran 扰动引起的 Ran 和 importin β 信号的变化,同时检查 MI 纺锤体动力学。我们发现,与之前研究中使用的 RanT24N 不同,RanT24N、T42A 双突变体抑制 RanGEF,而不干扰与导入蛋白 β 结合的货物,并破坏染色体分离中的 MI 纺锤体功能。 Ran 和 importin β 在微管组织中心 (MTOC) 合并和 MI 纺锤体组装中的作用得到了化学抑制剂 importazo 的使用进一步支持,其作用被 GTP 耐水解 RanQ69L 突变体部分挽救。这些结果表明 RanGTP 对于人类和小鼠的 MI 纺锤体组装和功能至关重要。
更新日期:2020-01-02
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