Studies of the structural rearrangements associated with the dynamic spliceosome

与动态剪接体相关的结构重排的研究

• 批准号: 9279192
• 负责人: Melanie Diane Ohi
• 金额: $ 13.36万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9279192
• 关键词: ATP phosphohydrolase; Affect; Alleles; Amino Acid Substitution; Animal Model; Biotin; Catalysis; Cell physiology; Chronic Lymphocytic Leukemia; Code; Complex; Computing Methodologies; Disease; Dissociation; Docking; Dysmyelopoietic Syndromes; Electrons; Epitopes; Excision; Fission Yeast; Genetic; Goals; Health; Imaging technology; Introns; Label; Lead; Ligase; Maps; Messenger RNA; Molecular; Molecular Conformation; Mutation; Nature; Nuclear; Patients; Process; Proteins; RNA; RNA Splicing; Reaction; Resolution; Role; Series; Small Nuclear Ribonucleoproteins; Spliceosome Assembly Pathway; Spliceosomes; Structure; Temperature; Transcript; U2 small nuclear RNA; Untranslated RNA; Work; detector; insight; loss of function; mRNA Precursor; molecular rearrangement; particle; protein complex; protein protein interaction; public health relevance; symposium; tumorigenesis

 DESCRIPTION (provided by applicant): The spliceosome is a dynamic macromolecular machine that catalyzes the excision of non-coding introns from pre-messenger RNAs (pre-mRNA) to form mature messages (mRNA), a process called pre- mRNA splicing. The ~3 MDa complex, composed of RNAs and proteins, assembles in a series of regulated steps from four small nuclear ribonucleoprotein subunits (snRNPs; U1, U2, U5, U4/U6) and numerous non- snRNP splicing factors. While the spliceosome is a fundamental cellular machine, its complex and dynamic nature has made obtaining structural information challenging. There are no structures of multi-snRNP (small nuclear ribonucleic particle) complexes at resolutions better than 30Å. The molecular organization of RNA and proteins within the spliceosome at any stage of the splicing reaction is not known and the global conformational changes that occur during the transitions from a pre- to post-splicing complex have not been characterized. The goal of this proposal is to define the conformational changes required for the transition from a pre- to post- activated spliceosome. In Aim 1 we will compare the structures of pre-activated, activated, and post-activated spliceosomes to map the global conformational changes required for pre-mRNA splicing. In Aim 2 we will define the molecular organization and map proximal protein-protein interaction networks of pre-activated, activated, and post-activated spliceosomes. In Aim 3 we will use structure/function studies to determine the role of the SF3 complex during catalytic activation and how this function is altered in Myelodysplastic Syndromes (MDS) and Chronic Lymphocytic Leukemia (CLL) patients that have mutations in this sub-complex. This work will provide direct insight into the conformational changes required for activation and define spliceosome organization during the transitions from a pre- to post-activated complex. Completion of these aims will significantly advance our understanding of the structure and organization of the spliceosome under conditions of both health and disease.

 描述（由适用提供）：剪接体是一种动态的大分子机器，它催化了来自通信剂RNA（PRE-MRNA）形成成熟消息（mRNA）的非编码内含子的惊喜，这是一个称为Pre-MRNA剪接的过程。由RNA和蛋白质组成的〜3 MDA复合物以一系列调节的步骤组成，从四个小的核核糖核蛋白亚基（SNRNP； U1，U2，U5，U4，U4/U6）和许多非SNRNP旋转因子组成。尽管剪接体是一种基本的蜂窝机，但其复杂而动态的性质使得获得了结构信息挑战。在分辨率大于30Å处的多SNRNP（小核带核颗粒）复合物的结构没有结构。在剪接体的任何阶段，RNA和蛋白质的分子组织尚不清楚，并且尚未表征从前至串联复合物过渡期间发生的全局构象变化。该提案的目的是定义从前至post激活的剪接体过渡所需的会议更改。在AIM 1中，我们将比较预激活，激活和后激活的剪接体的结构，以绘制前MRNA剪接所需的全局会议变化。在AIM 2中，我们将定义分子组织和映射代理蛋白 - 蛋白质 - 蛋白质相互作用网络的相互作用网络，该网络的激活，激活和激活后的剪接体。在AIM 3中，我们将使用结构/功能研究来确定SF3复合物在催化激活过程中的作用，以及该功能如何改变骨髓增生综合征（MDS）和慢性淋巴细胞性白血病（CLL）患者，这些患者在该亚副体积中具有突变。这项工作将直接洞悉激活所需的构象变化，并在从前激活的复合物到过渡期间定义剪接组组织。这些目标的完成将大大提高我们对健康和疾病条件下剪接体的结构和组织的理解。