Receptor-mediated substrate translocation through the nuclear pore complex without nucleotide triphosphate hydrolysis
Receptor-mediated substrate translocation through the nuclear pore complex without nucleotide triphosphate hydrolysis
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DOI：
10.1016/s0960-9822(99)80044-x
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发表时间：
1999-01-14
期刊：
影响因子：
9.2
通讯作者：
Mattaj, IW
中科院分区：
文献类型：
--
作者：
Englmeier, L;Olivo, JC;Mattaj, IW
背景：大分子物质在细胞核和细胞质之间的转运是一个能量依赖的过程。底物通过核孔复合物（NPC）跨核膜转运。易位需要输入蛋白β家族的核质转运受体，其与NPC相互作用，并且直接或通过接头与转运底物相互作用。虽然某些受体最近已被证明跨越NPC在一个能源独立的方式，通过NPC的底物-受体复合物的易位一般被认为是一个能源需求的step.Results：我们描述了一个体外系统，是基于透化细胞和支持核出口介导的亮氨酸丰富的核出口信号。在该系统中，输出依赖于外源性CRM 1/Exportin 1-一种核输出受体-GTdR Ran和三磷酸核苷酸（NTP），并进一步受到Ran结合蛋白1（RanBP 1）和核转运因子2（NTF 2）的刺激。出乎意料的是，不可水解的NTP类似物完全满足了单轮CRM 1介导的蛋白质底物跨NPC转运的NTP要求。同样，单一的转运蛋白介导的核蛋白进口事件不需要水解的NTPs和发生在没有的RAN GTsald.Conclusions：我们的数据表明，与预期和先前的结论相反，在任何方向的基板受体复合物的易位在NPC中发生在没有NTP水解，因此是能量独立的。在细胞质中受体再循环的步骤中提供驱动底物逆浓度梯度转运所需的能量。
Background: The transport of macromolecules between the nucleus and cytoplasm is an energy-dependent process. Substrates are translocated across the nuclear envelope through nuclear pore complexes (NPCs). Translocation requires nucleocytoplasmic transport receptors of the importin beta family, which interact both with the NPC and, either directly or via an adaptor, with the transport substrate. Although certain receptors have recently been shown to cross the NPC in an energy-independent manner, translocation of substrate-receptor complexes through the NPC has generally been regarded as an energy-requiring step.Results: We describe an in vitro system that is based on permeabilised cells and supports nuclear export mediated by leucine-rich nuclear export signals. In this system, export is dependent on exogenous CRM1/Exportin1 - a nuclear export receptor - the GTPase Ran and nucleotide triphosphates (NTPs), and is further stimulated by Ran-binding protein 1 (RanBP1) and nuclear transport factor 2 (NTF2). Unexpectedly, non-hydrolysable NTP analogues completely satisfy the NTP requirements for a single-round of CRM1-mediated translocation of protein substrates across the NPC. Similarly, single transportin-mediated nuclear protein import events are shown not to require hydrolysable NTPs and to occur in the absence of the Ran GTPase.Conclusions: Our data show that, contrary to expectation and prior conclusions, the translocation of substrate-receptor complexes across the NPC in either direction occurs in the absence of NTP hydrolysis and is thus energy independent. The energy needed to drive substrate transport against a concentration gradient is supplied at the step of receptor recycling in the cytoplasm.