Structures and mechanisms of nuclear import and export

描述（由申请人提供）：核粘蛋白通过识别不同的核定位或输出信号（NLSs或NESs）介导核细胞质运输。虽然人类已知有19个Kaps，但目前只知道Kap¿/Kap¿1、Kap¿2（或运输蛋白）和CRM1的三类信号。大多数其他的Kaps识别出不同的序列，这些序列掩盖了它们信号的身份。本提案描述了进口系统Kap¿2和Imp5/Kap121以及出口系统Kap CRM1的结构和生化分析。Kap¿2通过其PY-NLSs介导RNA结合蛋白的核输入。我们对PY-NLS的发现和理解有助于发现导致肌萎缩性侧索硬化症（ALS）亚群的缺陷性PY-NLS。在此，我们提出了解Kap¿2在ALS蛋白错定位和聚集中的作用。在另一个目标中，我们将把我们用来发现PY-NLS的结构、生化和生物信息学相结合的方法应用于Imp5/Kap121p途径，目的是发现一组特征来统一Kaps识别的不同序列，从而揭示一类新的NLS。最后，我们将研究介导数百种蛋白质核输出的CRM1；其中大多数是使用小分子抑制剂Leptomycin B鉴定的。本提案旨在确定CRM1如何介导Leptomycin B偶联以抑制核输出的机制。

DESCRIPTION (provided by applicant): Karyopherin¿ proteins mediate nucleocytoplasmic transport through recognition of distinct nuclear localization or export signals (NLSs or NESs). Although there are 19 known Kaps in humans, only three classes of signals for Kap¿/Kap¿1, Kap¿2 (or Transportin) and CRM1, respectively, are known at this time. Most other Kaps recognize diverse sequences that occlude the identity of their signals. This proposal describes structural and biochemical analyses of import systems Kap¿2 and Imp5/Kap121 as well as the export-Kap CRM1. Kap¿2 mediates nuclear import of RNA binding proteins through their PY-NLSs. Our discovery and understanding of the PY-NLS contributed to the discovery of a defective PY-NLS that causes a subset of amyotrophic lateral sclerosis (ALS). Here, we propose to understand the role of Kap¿2 in protein mislocalization and aggregation in ALS. In another aim, we will apply the combined structural, biochemical and bioinformatics approach that we used to discover the PY-NLS to the Imp5/Kap121p pathway with the goal of discovering a set of characteristics to unify the diverse sequences recognized by the Kaps, hence revealing a new class of NLS. Finally, we will study CRM1, which mediates nuclear export of hundreds of proteins; most of them identified using the small molecule inhibitor Leptomycin B. This proposal aims to determine the mechanism of how CRM1 mediates conjugation of leptomycin B to inhibit nuclear export.