Characterization of the mammalian mRNA-3'-end processing complex

哺乳动物 mRNA-3 末端加工复合物的表征

• 批准号: 8963877
• 负责人: Yongsheng Shi
• 金额: $ 30.2万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-15 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8963877
• 关键词: Amino Acids; Beryllium; Binding; Complex; Data; Development; Disease; Elements; Foundations; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genetic Translation; Goals; In Vitro; Integration Host Factors; Maintenance; Malignant Neoplasms; Mediating; Messenger RNA; Metabolism; Modeling; Molecular; Neuromuscular Diseases; Nuclear; Pattern; Play; Polyadenylation; Process; Protein Isoforms; Proteins; Publishing; RNA; RNA Binding; RNA Splicing; RNA-Binding Proteins; RNA-Protein Interaction; Regulation; Research; Role; Seminal; Site; Specificity; Structure; Translations; Virus Diseases; base; clinically relevant; human disease; in vivo; influenzavirus; insight; mRNA Stability; novel; novel therapeutics; prevent; public health relevance

 DESCRIPTION (provided by applicant): Project Summary: The long-term goal of this proposal is to understand, in detail, the mechanisms of mammalian mRNA 3' processing and its regulation. mRNA 3'-end formation, typically involving an endonucleolytic cleavage followed by polyadenylation, is an essential step of eukaryotic gene expression and it significantly impacts many aspects of RNA metabolism, including mRNA stability and translation. In addition, the majority of eukaryotic genes produce multiple mRNA isoforms with distinct 3' ends through alternative polyadenylation (APA). Recent studies have revealed that APA is highly regulated in development and plays an important role in post-transcriptional gene regulation. Aberrant APA patterns have been associated with a wide range of diseases, from cancer to neuromuscular disorders. As such, a central question in the mRNA 3' processing field has been how polyadenylation sites (PAS) are recognized and how PAS selection can be regulated. The majority of mammalian PAS contain an AAUAAA hexamer and a U/UG-rich downstream element (DSE). According to the current model in the field, these key cis-elements are specifically recognized by CPSF160 of the CPSF complex and CstF64 of the CstF complex respectively. However, our published and preliminary data have challenged this model in several key aspects: 1) we have recently shown that the CPSF subunits CPSF30 and Wdr33, but not CPSF160 as was generally believed, directly bind to AAUAAA; 2) we have provided evidence that maintenance of the CPSF-RNA interaction specificity requires a novel proofreading factor(s) (see preliminary data); 3) we have demonstrated that the general 3' processing factor CstF64 in fact only binds to a subset of mammalian PAS, and we provided evidence that recognition of PAS with different sequence features requires distinct RNA-binding protein(s) (see preliminary data). These observations have not only significantly changed the current model for PAS recognition, but also revealed much greater complexity in mRNA 3' processing than previously appreciated. Here we propose to better define the molecular mechanisms of mammalian PAS recognition through an in-depth characterization of the key protein-RNA interactions involved. Accomplishing the proposed research will provide seminal insights into the fundamental mechanisms of mammalian mRNA 3' processing and its regulation. As aberrant PAS selection has been associated with a broad spectrum of human diseases, a better understanding of the mechanisms for PAS recognition may provide the foundation for the development of new therapeutics for these diseases.

 描述(由申请人提供)： 项目摘要：这项建议的长期目标是详细了解哺乳动物信使核糖核酸3‘加工及其调控的机制。MRNA3‘端的形成是真核基因表达的重要步骤，它对RNA代谢的许多方面都有重要影响，包括mRNA的稳定性和翻译。此外，大多数真核基因通过交替多聚腺苷酸化(APA)产生具有不同3‘端的多种mRNA异构体。最近的研究表明，APA在发育过程中受到高度调控，并在转录后基因调控中发挥重要作用。APA模式的异常与多种疾病有关，从癌症到神经肌肉疾病。因此，mRNA3‘加工领域的一个中心问题是如何识别多聚腺苷酸化位点(PA)以及如何调控PAS选择。大多数哺乳动物PA含有AAUAAA六聚体和富含U/UG的下游元件(DSE)。根据目前的模型，这些关键的顺式元件分别被CPSF复合体的CPSF160和CstF复合体的CstF64特异性识别。然而，我们已发表的和初步的数据在几个关键方面对该模型提出了质疑：1)我们最近发现CPSF亚单位CPSF30和Wdr33，而不是通常认为的CPSF160直接与AAUAAA结合；2)我们提供的证据表明，保持CPSF-RNA的相互作用专一性需要一个新的校对因子(S)(见初步数据)；3)我们已经证明一般的3‘处理因子CstF64实际上只与哺乳动物PA的一个亚组结合，并且我们提供的证据表明识别具有不同序列特征的PA需要不同的RNA结合蛋白(见初步数据)(见初步数据)。这些观察结果不仅显著改变了目前PAS识别的模型，而且揭示了mRNA3‘处理的复杂性比以前所认识的要大得多。在这里，我们建议通过深入描述所涉及的关键蛋白质-RNA相互作用来更好地定义哺乳动物PAS识别的分子机制。这项研究的完成将为哺乳动物mRNA3‘处理及其调控的基本机制提供开创性的见解。由于PAS的异常选择与人类多种疾病有关，对PAS识别机制的更好理解可能为这些疾病的新疗法的开发提供基础。