KARYOPHERIN X, A NOVEL TRANSPORT RECEPTOR

蛋白质和RNA在细胞质和细胞核之间的转运是由一个可溶性转运受体家族介导的，迄今为止在哺乳动物中仅鉴定出其中的九种。 了解这些因素的重要性，强调了相关的研究结果，不正确的定位关键调控蛋白质的疾病状态，如致癌。 此外，几种转运蛋白在病毒的生命周期和发病机制中起着不可或缺的作用（例如，HIV）。 拟议的研究计划是基于鉴定和克隆一种新的人类转运受体，核转运蛋白X。 其目标是对核转运蛋白X进行生物化学表征，鉴定其货物并绘制其识别并结合货物序列内的基序。 为了实现这些目标，将追求两个具体目标。 第一个目标是确定这种核转运蛋白是否具有定义输入素β样转运受体家族成员的生化特性。 第二个目标是通过鉴定其货物并显示其介导该货物穿过核孔的易位来验证该蛋白是真正的转运蛋白的假设。 目的1实验测试Ran（一种小的GT3）结合的转运蛋白、影响Ran核苷酸状态的因子的抑制以及与核孔蛋白的相互作用。目标2的实验方法包括使用固定化转运蛋白和细胞裂解物的下拉测定以及透化细胞中的体外转运测定。 从长远来看，工作将集中在定位定位序列，分类货物的核粒蛋白X介导的运输。

The translocation of proteins and RNA between the cytoplasm and nucleus is mediated by a family of soluble transport receptors, of which only nine have so far been identified in mammals. The importance of understanding these factors is underscored by findings that correlate the improper localization of key regulatory proteins to disease states such as carcinogenesis. In addition, several transporters play integral roles in the life cycle and pathogenesis of viruses (e.g., HIV). The proposed research plan is based on the identification and cloning of a novel, human transport receptor, karyopherin X. The objectives are to biochemically characterize karyopherin X, identify its cargo and map the motif it recognizes and binds to within the cargo sequence. To realize these goals, two specific aims will be pursued. Aim number 1 is to determine if this karyopherin shares the biochemical properties that define the members of the importin-beta-like transport receptor family. Aim number 2 is to test the hypothesis that this protein is a bona fide transporter, by identifying its cargo and showing that it mediates the translocation of this cargo across the nuclear pore. Aim number 1 experiments test the transporter for Ran (a small GTPase) binding, inhibition of the factors that influence the nucleotide state of Ran, and interactions with nuclear pore proteins. Experimental approaches for Aim number 2 include pulldown assays with immobilized transporter and cell lysates and in vitro transport assays in permeabilized cells. In the longer term, efforts will focus on mapping the targeting sequence(s) that sorts cargo for karyopherin X-mediated transport.