权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Analysis of transport mechanism and regulation through the recycling of transport factors.

通过运输要素的循环利用分析运输机制和调控。

基本信息

批准号：
13480240
负责人：
IMAMOTO Naoko
金额：
$ 9.73万
依托单位：
RIKEN (2002)National Institute of Genetics (2001)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13480240/
关键词：
nuclear transport importin β CAS small GTPase Ran importin α nuclear pore complex 70kDa heat-shock cognate protein (hsc70)β-catenin 核内輸送担体importinβ 核外輸送担体CAS 核膜孔複合体 p10 NTF2 hsc70 輸送担体リサイクリング

项目摘要

1.We have initially examined nuclear import and export of importin β in living cells, and found that its nuclear export depends on the presence of energy sources, whereas its import occurs in an energy-independent manner. Analysis in digitonin-permeabilized cell-free system and through the fractionation studies of cytosol, we found that nuclear export of importin β is stimulated not only by small GTPase Ran but also by 70kDa heat-shock cognate protein (hsc70). Point mutated hsc70 lacking ATPase did not stimulate nuclear export of importin β. Our results indicate that additional factor is involved in recycling of importin β and the stimulation is dependent on ATPase activity. 2.β-catenin is an example of a typical molecule that can be bi-directionally translocated through nuclear pore complexes (NPC) on its own in a facilitated manner. We examined the nuclear import and export of β-catenin, and showed that region of β-catenin required for import and export substantially overlapped. However, different nucleocytoplasmic shuttling molecules such as importin β and GAS, differentially blocked nuclear import and export of β-catenin. The different effect was also observed under the condition that induces recycling of transport factor. These data indicates that β-catenin shows an overlapping sequence requirement for its import and export, but that bi-directional movement through the NPC proceeds through distinct molecular interactions. 3.We found that importin α/β mediated nuclear import pathway is down-regulated in response to heat-shock, which was caused by nuclear retention and recycling inhibition of importin α. Such regulation of transport suggests a new regulatory mechanism for the adaptation of cells to environmental changes, such as heat-shock.

1.我们初步研究了活细胞中importin β的核输入和输出，发现其核输出依赖于能源的存在，而其输入则以与能量无关的方式发生。在洋地黄皂苷通透的无细胞系统中进行分析并通过细胞质的分级分离研究，我们发现importin β的核输出不仅受到小GTPase Ran的刺激，而且还受到70kDa热休克同源蛋白(hsc70)的刺激。缺乏 ATP 酶的点突变 hsc70 不会刺激输入蛋白 β 的核输出。我们的结果表明，输入蛋白 β 的再循环涉及其他因素，并且刺激取决于 ATP 酶活性。 2.β-连环蛋白是典型分子的一个例子，它可以通过核孔复合物（NPC）以便利的方式自行双向易位。我们检查了β-连环蛋白的核进出口，结果表明进出口所需的β-连环蛋白区域基本重叠。然而，不同的核质穿梭分子，例如输入蛋白β和GAS，差异性地阻断β-连环蛋白的核输入和输出。在诱导转运因子再循环的条件下也观察到不同的效果。这些数据表明，β-连环蛋白的输入和输出显示出重叠的序列要求，但通过 NPC 的双向运动是通过不同的分子相互作用进行的。 3.我们发现importin α/β介导的核输入途径在热激反应中下调，这是由importin α的核滞留和再循环抑制引起的。这种运输调节表明细胞适应环境变化（例如热休克）的新调节机制。