Yamada M, Tachibana T, Imamoto N, Yoneda Y
Department of Anatomy and Cell Biology Osaka University Medical School 2-2 Yamada-oka Suita Osaka 565-0871 Japan.
Curr Biol. 1998 Dec 3;8(24):1339-42. doi: 10.1016/s0960-9822(07)00566-0.
The cytosolic nuclear transport factor p10/NTF2 is required for the translocation of karyophilic molecules through nuclear pores [1] [2] [3], and the small GTPase Ran is a key regulator of protein transport between the nucleus and cytoplasm [4] [5]. It has been reported that p10/NTF2 interacts directly and specifically with Ran-GDP but not with Ran-GTP [6]. The precise role(s) of p10/NTF2 in the Ran GTP/GDP cycle are thus far unclear, however. In this study, we show that mammalian p10/NTF2 dramatically inhibits the dissociation of [3H]GDP from Ran and the binding of [35S]GTPgammaS to Ran following the dissociation of non-radioactive GDP by RCC1, the only known mammalian guanine nucleotide exchange factor for Ran (Ran-GEF) [7]. In contrast, the dissociation of [35S]GTP gamma S from Ran, which was also catalyzed by RCC1, was not affected by p10/NTF2. Furthermore, the activities of wild-type p10/NTF2 and the mutant forms M84T and D92G in an assay of nuclear protein import in a digitonin-permeabilized cell-free system correlated with their level of inhibition of the dissociation of nucleotide from Ran-GDP. These results suggest that p10/NTF2 acts as a GDP dissociation inhibitor for Ran (Ran-GDI), thereby coordinating the Ran-dependent reactions that underlie nuclear protein import.
胞质核转运因子p10/NTF2是亲核分子通过核孔进行转运所必需的[1][2][3]，而小GTP酶Ran是细胞核与细胞质之间蛋白质运输的关键调节因子[4][5]。据报道，p10/NTF2直接且特异性地与Ran-GDP相互作用，而不与Ran-GTP相互作用[6]。然而，到目前为止，p10/NTF2在Ran GTP/GDP循环中的确切作用尚不清楚。在本研究中，我们发现，在唯一已知的哺乳动物Ran鸟嘌呤核苷酸交换因子（Ran-GEF）RCC1促使非放射性GDP解离后，哺乳动物p10/NTF2能显著抑制[3H]GDP从Ran上的解离以及[35S]GTPγS与Ran的结合[7]。相比之下，同样由RCC1催化的[35S]GTPγS从Ran上的解离不受p10/NTF2的影响。此外，在洋地黄皂苷通透的无细胞体系中进行的核蛋白输入测定中，野生型p10/NTF2以及突变形式M84T和D92G的活性与它们对Ran-GDP核苷酸解离的抑制水平相关。这些结果表明，p10/NTF2作为Ran的GDP解离抑制剂（Ran-GDI），从而协调了核蛋白输入所依赖的Ran相关反应。
