Translocation of cyclin B1 to the nucleus at prophase requires a phosphorylation-dependent nuclear import signal
Translocation of cyclin B1 to the nucleus at prophase requires a phosphorylation-dependent nuclear import signal
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DOI：
10.1016/s0960-9822(99)80308-x
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发表时间：
1999-07-01
期刊：
影响因子：
9.2
通讯作者：
Pines, J
中科院分区：
文献类型：
--
作者：
Hagting, A;Jackman, M;Pines, J
背景：在 M 期，细胞周期蛋白 B1 在细胞质保留序列 (CRS) 中被磷酸化，这是核输出所需的。在间期期间，细胞周期蛋白 B1 在细胞核和细胞质之间穿梭，因为组成型核输入被快速核输出所抵消。在 M 期，细胞周期蛋白 B 在其磷酸化的同时迅速进入细胞核，这一整体运动可能仅仅是由于其核输出的减少而引起的。然而，有丝分裂中细胞周期蛋白 B1 易位是否需要 CRS 磷酸化以及核输出减少是否足以解释其快速重新定位的问题尚未得到解决。 结果：我们使用两种形式的绿色荧光蛋白同时分析有丝分裂中野生型细胞周期蛋白 B1 和细胞周期蛋白 B1 磷酸化突变体的易位，并将其与体外核输入测定相关联。我们发现，在核膜破裂前大约 10 分钟，细胞周期蛋白 B1 迅速易位到细胞核中，并且这种运动需要 CRS 磷酸化位点。不能被磷酸化的细胞周期蛋白B1突变体在野生型蛋白之后进入细胞核。 CRS 的磷酸化会产生一个核输入信号，该信号在体外和体内增强细胞周期蛋白 B1 的输入，其方式与先前描述的由输入蛋白 β 介导的细胞周期蛋白 B1 的输入不同。结论：我们表明，人细胞周期蛋白 B1 的磷酸化是其在前期末期快速易位到细胞核所必需的。磷酸化通过产生核输入信号来增强细胞周期蛋白 B1 的核输入。因此，CRS 的磷酸化是有丝分裂控制的关键步骤。
Background: At M phase, cyclin B1 is phosphorylated in the cytoplasmic retention sequence (CRS), which is required for nuclear export. During interphase, cyclin B1 shuttles between the nucleus and the cytoplasm because constitutive nuclear import is counteracted by rapid nuclear export. In M phase, cyclin B moves rapidly into the nucleus coincident with its phosphorylation, an overall movement that might be caused simply by a decrease in its nuclear export. However, the questions of whether CRS phosphorylation is required for cyclin B1 translocation in mitosis and whether a reduction in nuclear export is sufficient to explain its rapid relocalisation have not been addressed.Results: We have used two forms of green fluorescent protein to analyse simultaneously the translocation of wild-type cyclin B1 and a phosphorylation mutant of cyclin B1 in mitosis, and correlated this with an in vitro nuclear import assay. We show that cyclin B1 rapidly translocates into the nucleus approximately 10 minutes before breakdown of the nuclear envelope, and that this movement requires the CRS phosphorylation sites. A cyclin B1 mutant that cannot be phosphorylated enters the nucleus after the wild-type protein. Phosphorylation of the CRS creates a nuclear import signal that enhances cyclin B1 import in vitro and in vivo, in a manner distinct from the previously described import of cyclin B1 mediated by importin beta.Conclusions: We show that phosphorylation of human cyclin B1 is required for its rapid translocation to the nucleus towards the end of prophase. Phosphorylation enhances cyclin B1 nuclear import by creating a nuclear import signal. The phosphorylation of the CRS is therefore a critical step in the control of mitosis.