ABSTRACT

The Ran GTPase plays critical roles in multiple cellular processes including interphase nucleocytoplasmic transport and mitotic spindle assembly. During mitosis in mammalian cells, GTP-bound Ran (Ran-GTP) is concentrated near mitotic chromatin while GDP-bound Ran (Ran-GDP) is more abundant distal to chromosomes. This pattern spatially controls spindle formation because Ran-GTP locally releases spindle assembly factors (SAFs), such as Hepatoma Up-Regulated Protein (HURP), from inhibitory interactions near chromosomes. Regulator of Chromatin Condensation 1 (RCC1) is Ran’s chromatin-bound exchange factor, and RanBP1 is a conserved Ran-GTP-binding protein that has been implicated as a mitotic regulator of RCC1 in embryonic systems. Here, we show that RanBP1 controls mitotic RCC1 dynamics in human somatic tissue culture cells. In addition, we observed the re-localization of HURP in metaphase cells after RanBP1 degradation, consistent with the idea that altered RCC1 dynamics functionally modulate SAF activities. Together, our findings reveal an important mitotic role for RanBP1 in human somatic cells, controlling the spatial distribution and magnitude of mitotic Ran-GTP production and thereby ensuring the accurate execution of Ran-dependent mitotic events.

Abbreviations

AID: Auxin-induced degron; FLIP: Fluorescence loss in photobleaching; FRAP: Fluorescence recovery after photobleaching; GDP: guanosine diphosphate; GTP: guanosine triphosphate; HURP: Hepatoma Up-Regulated Protein; NE: nuclear envelope; NEBD: Nuclear Envelope Breakdown; RanBP1: Ran-binding protein 1; RanGAP1: Ran GTPase-Activating Protein 1; RCC1: Regulator of Chromatin Condensation 1; RRR complex: RCC1/Ran/RanBP1 heterotrimeric complex; SAF: Spindle Assembly Factor; TIR1: Transport Inhibitor Response 1 protein; XEE: Xenopus egg extract.

中文翻译：

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RanBP1 通过调节有丝分裂的 RCC1 动力学来控制哺乳动物细胞中的 Ran 通路。

摘要

Ran GTPase 在多个细胞过程中起着关键作用，包括相间核细胞质运输和有丝分裂纺锤体组装。在哺乳动物细胞的有丝分裂过程中，GTP 结合的 Ran (Ran-GTP) 集中在有丝分裂染色质附近，而 GDP 结合的 Ran (Ran-GDP) 在染色体远端更丰富。这种模式在空间上控制纺锤体的形成，因为 Ran-GTP 从染色体附近的抑制相互作用中局部释放纺锤体组装因子 (SAF)，例如肝癌上调蛋白 (HURP)。染色质凝聚调节因子 1 (RCC1) 是 Ran 的染色质结合交换因子，RanBP1 是一种保守的 Ran-GTP 结合蛋白，在胚胎系统中被认为是 RCC1 的有丝分裂调节因子。在这里，我们表明 RanBP1 控制人类体细胞组织培养细胞中的有丝分裂 RCC1 动力学。此外，我们观察到 HURP 在 RanBP1 降解后中期细胞中的重新定位，这与改变 RCC1 动力学在功能上调节 SAF 活动的想法一致。总之，我们的研究结果揭示了 RanBP1 在人类体细胞中的重要有丝分裂作用，控制有丝分裂 Ran-GTP 产生的空间分布和大小，从而确保 Ran 依赖性有丝分裂事件的准确执行。

缩写

AID：生长素诱导的降解；FLIP：光漂白中的荧光损失；FRAP：光漂白后的荧光恢复；GDP：鸟苷二磷酸；GTP：三磷酸鸟苷；HURP：肝癌上调蛋白；NE：核膜；NEBD：核包络分解；RanBP1：Ran结合蛋白1；RanGAP1：Ran GTPase 激活蛋白 1；RCC1：染色质凝聚调节剂1；RRR复合体：RCC1/Ran/RanBP1异源三聚体复合体；SAF：主轴装配系数；TIR1：运输抑制剂反应 1 蛋白；XEE：非洲爪蟾卵提取物。