Messenger RNA transport across the nuclear pore complex

信使 RNA 穿过核孔复合体的运输

• 批准号: 7118803
• 负责人: KARSTEN WEIS
• 金额: $ 27.16万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-30 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7118803
• 关键词: RNA binding protein; Saccharomyces cerevisiae; biological signal transduction; fungal genetics; guanosinetriphosphatases; helicase; in situ hybridization; intracellular transport; messenger RNA; molecular cloning; nuclear membrane; nucleic acid sequence; protein structure function; protein transport; site directed mutagenesis; transport proteins

DESCRIPTION (provided by applicant): Transport of macromolecules between the nucleus and the cytoplasm is an essential cellular process in all eukaryotes. The maintenance and decoding of the eukaryotic genome, and the dynamic state of the eukaryotic transcriptome and proteome relies on the compartmentalization and exchange of a large number of proteins and RNAs across the nuclear envelope. Furthermore, it is well documented that the regulation of nucleocytoplasmic transport provides an important mechanism by which signal transduction pathways and developmental stimuli control differential gene expression in eukaryotes. In addition, many viruses target components of the cellular nuclear transport machinery, and exploit or modify them to promote viral propagation. Therefore, a better understanding of the molecular machinery that mediates nucleocytoplasmic transport is essential both for understanding fundamental cellular processes and the development of novel anti-viral therapies. Despite the critical importance of messenger RNA (mRNA) export for eukaryotic gene expression, many aspects of the packaging, processing, and transport of mRNA-containing ribonucleoprotein particles (RNPs) from the nucleus have not yet been elucidated. The long-term objective of the research program described in this proposal is to understand the molecular pathway by which mRNAs are targeted to and translocated across the nuclear envelope. Export of mRNA appears to be mediated by multiple soluble protein factors that specifically bind to mRNA in the nucleus but release their cargo in the cytoplasm upon translocation through the nuclear pore complex. This spatial regulation of cargo binding and release is important for the transport of mRNAs but remains poorly understood at the mechanistic level. Moreover, evidence obtained in our laboratory and others indicates an important role for soluble, inositol polyphosphates in mRNA export, but the target(s) of these effectors have not been identified. Thus, we specifically propose: (1) to characterize the role of the major poly (A)-binding protein Pabl in mRNA maturation and export; (2) to identify the function and the targets of soluble inositol polyphosphates in mRNA export; and, (3) to trap, isolate, and characterize intermediates of the mRNA export pathway and to determine how mRNA export complexes are disassembled in the cytoplasm. The proposed experiments take advantage of the proteomic and genomic tools available in the yeast Saccharomyces cerevisiae and employ a combination of innovative biochemical, genetic and cell biological approaches to address these three specific aims. Because mRNA transport is a highly conserved process, the mechanistic insights obtained from these studies will be directly relevant to all eukaryotes, including humans.

描述（由申请人提供）：细胞核和细胞质之间大分子的运输是所有真核生物中必不可少的细胞过程。真核基因组的维持和解码以及真核转录组和蛋白质组的动态状态依赖于整个核包膜上大量蛋白质和RNA的分隔和交换。此外，有充分的文献证明，核质转运的调节提供了一种重要的机制，通过该机制，信号转导途径和发育刺激控制真核生物中的差异基因表达。此外，许多病毒靶向细胞核转运机制的成分，并利用或修改它们以促进病毒传播。因此，更好地理解介导核细胞质转运的分子机制对于理解基本细胞过程和新型抗病毒疗法的发展至关重要。尽管信使RNA（mRNA）对真核基因表达的重要性至关重要，但尚未阐明含核核糖核蛋白颗粒（RNP）包装，加工和运输的许多方面。该提案中描述的研究计划的长期目标是了解mRNA靶向和易位的分子途径。 mRNA的导出似乎是由多种可溶性蛋白因子介导的，这些因子特异性结合了细胞核中的mRNA，但通过核孔复合物易位后将其货物释放在细胞质中。货物结合和释放的这种空间调节对于mRNA的运输至关重要，但在机械水平上仍然了解得很糟糕。此外，在我们的实验室和其他方面获得的证据表明，在mRNA输出中可溶性，肌醇多磷酸盐的重要作用，但尚未确定这些效应子的靶标。因此，我们特别提出：（1）表征主要的聚（A）结合蛋白PABL在mRNA成熟和导出中的作用； （2）确定mRNA输出中可溶性肌醇多磷酸盐的功能和靶； （3）要捕获，分离和表征mRNA输出途径的中间体，并确定mRNA输出复合物如何在细胞质中解散。提出的实验利用了酿酒酵母中可用的蛋白质组学和基因组工具，并采用了创新的生化，遗传和细胞生物学方法的组合来解决这三个特定目标。由于mRNA转运是一个高度保守的过程，因此从这些研究中获得的机械见解将与包括人类在内的所有真核生物直接相关。