MECHANISM OF MACROMOLECULAR EXPORT FROM THE NUCLEUS

描述（改编自申请人的摘要）：RNA 和 蛋白质从细胞核进入细胞质是细胞的基本物质 过程和控制真核基因表达的关键步骤。的 管理这些运输事件的机制仍然知之甚少 但爪蟾卵母细胞的动力学竞争研究表明 不同的 RNA 类别，包括 mRNA、snRNA、tRNA 和 rRNA 使用不同的 出口途径。因为大多数（如果不是全部）RNA 都与蛋白质相关 在细胞核中，有人认为 RNA 输出事件是由 细胞核中的蛋白质表明 RNA 输出事件是 由含有适当核输出信号（NES）的蛋白质介导。 这是由小可转移信号的识别支持的 导致其从细胞核快速、主动输出的蛋白质。的 拟议研究的长期目标是表征不同的RNA 和酿酒酵母中的蛋白质输出途径并了解 分子水平上的核输出机制。更具体地说， 提出了三个具体目标：（1）P.I.将表征 核输出因子1（Xpo1p/Crm1p）的输出及其在mRNA中的作用 出口。将使用多种生化和遗传学方法 确定与 Xpo1p 相互作用的因素。 GTPase Ran 的作用 调节 Xpo1p 与其他细胞因子的相互作用将是 检查了介导 Xpo10,s 在 mRNA 输出中的作用的接头 将被表征。 (2) NES介导的核蛋白输出 机械将被解剖。将探索遗传方法 确定 NES 蛋白质输出机制的其他组件。这个 输出途径将被用作模型系统来理解分子 大分子如何通过核孔运输的细节 复合物进入细胞质。 (3) 最后，P.I.将确定是否 酵母中存在不同的 RNA 输出途径。固定和原位标记 活细胞将用于表征不同 RNA 的输出 几个突变体背景中的类。如果不同的路径是 确定后，调解和规范这些出口事件的因素将是 确定。

DESCRIPTION (adapted from the applicant's abstract): Export of RNA and protein from the nucleus into the cytoplasm is a fundamental cellular process and a key step in the control of eukaryotic gene expression. The mechanisms governing these transport events are still poorly understood but kinetic competition studies in Xenopus oocytes have indicated that different RNA classes, including mRNA, snRNA, tRNA and rRNA use distinct export pathways. Since most, if not all RNAs, are associated with proteins in the nucleus it was suggested that RNA export events are mediated by proteins in the nucleus it was suggested that RNA export events are mediated by proteins containing appropriate nuclear export signals (NESs). This is supported by the identification of small transferable signals in proteins that cause their rapid and active export from the nucleus. The long-term goal of the proposed research is to characterize different RNA and protein export pathways in the yeast S. cerevisiae and to understand the nuclear export mechanisms at a molecular level. More specifically, three specific aims are proposed: (1) The P.I. will characterize the nuclear export factor export in 1 (Xpo1p/Crm1p) and its role in mRNA export. A variety of biochemical and genetic approaches will be used to identify factors which interact with Xpo1p. The role of the GTPase Ran in regulating the interaction of Xpo1p with other cellular factors will be examined and the adapter(s) that mediate(s) Xpo10,s role in mRNA export will be characterized. (2) The NES-mediated nuclear protein export machinery will be dissected. Genetic approaches will be explored to identify additional components of the NES-protein export machinery. This export pathway will be used as a model system to understand the molecular details of how macromolecules are transported through the nuclear pore complex into the cytoplasm. (3) Finally, the P.I. will determine whether distinct RNA export pathways exist in yeast. In situ labeling in fixed and live cells will be used to characterize the export of the different RNA classes in several mutant backgrounds. If distinct pathways are identified, factors that mediate and regulate these export events will be identified.