Methylation of Sm proteins - Roles in snRNP Biogenesis and germline specification

Sm 蛋白的甲基化 - snRNP 生物发生和种系规范中的作用

• 批准号: 7471439
• 负责人: Graydon Gonsalvez
• 金额: $ 5.29万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-01 至 2009-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7471439
• 关键词: Address; Affinity; Animals; Arginine; Binding; Biogenesis; Biological Assay; Biological Models; C-terminal; Caenorhabditis elegans; Cell Culture Techniques; Cell Differentiation process; Cells; Childhood; Class; Complex; Cytoplasmic Granules; Defect; Development; Disease; Disruption; Drosophila genus; Enzymes; Etiology; Eukaryota; Eukaryotic Cell; Event; Excision; Exons; Failure; Gap Junctions; Genes; Germ Cells; Goals; Hela Cells; Hereditary Disease; Human; Hybrids; Immunoelectron Microscopy; Introns; Kinetics; Life; Light; Localized; Mammalian Cell; Mammals; Mediating; Messenger RNA; Methylation; Modification; Molecular; Motor Neurons; Mus; Mutation; Names; Neurodegenerative Disorders; Nexus (resin cement); Orthologous Gene; Ovary; PRDM1 gene; Pathology; Pathway interactions; Patients; Phenocopy; Phenotype; Post-Translational Protein Processing; Process; Protein-Arginine N-Methyltransferase; Proteins; Protocols documentation; Purpose; RNA; RNA Interference; RNA Splicing; Ribonucleoproteins; Role; Role playing therapy; SMN protein (spinal muscular atrophy); Sampling; Severity of illness; Small Nuclear RNA; Small Nuclear Ribonucleoproteins; Specific qualifier value; Spinal Muscular Atrophy; Spliceosomes; Structure; Tail; Techniques; Testing; Thinking; To specify; Work; cellular imaging; dimethylarginine; early childhood; fly; in vivo; insight; knock-down; light microscopy; mortality; mutant; research study; snRNP Biogenesis; snRNP Structural Core Protein; tool

DESCRIPTION (provided by applicant): Sm class small nuclear ribonucleoproteins (snRNPs) are core components of the spliceosome and are required for splicing in vivo. In addition, Sm proteins appear to have a more ancestral role in specification of the germline. The overall goal of this project is to understand the mechanism by which Sm proteins are assembled into snRNPs for the purpose of splicing or into specialized ribonucleoproteins (RNPs) required to establish germ cell fate. At the nexus of these two seemingly disparate pathways is the PRMT5 complex. The biogenesis of snRNPs is highly orchestrated, requiring several essential co-factors. In particular, cytoplasmic snRNP assembly relies on two heteromeric complexes, the PRMT5 complex and the SMN complex. Importantly, mutations that reduce the level of SMN protein are correlated with a neurodegenerative disease known as Spinal Muscular Atrophy (SMA). Patients with the disease often die very early in childhood. The molecular etiology of SMA is unknown, however, perturbation of snRNP biogenesis is thought to be a major contributing factor. A detailed understanding of the mechanism of snRNP assembly will shed light on the disease process. The PRMT5 complex is responsible for symmetrically dimethylating Sm proteins, and by virtue of this activity, is thought to cooperate with the SMN complex in mediating efficient snRNP assembly. However, an in vivo examination of this process is currently lacking. Furthermore, PRMT5 associates with BLIMP1 and is required for germ cell development in the mouse. Disruption of PRMT5 activity could thus impinge on two distinct but separable pathways - snRNP biogenesis and germ cell development. To gain mechanistic insight into these processes we have developed in vivo protocols making use of Drosophila as a model system as well as mammalian cell culture techniques. Aim 1 explores the requirement for Sm protein methylation in snRNP biogenesis. Since SMN binds with a higher affinity to methylated Sm proteins, this modification may act as a modifier of the SMA phenotype. Specifically, lack of Sm protein methylation in mammals may phenocopy SMA. Alternatively, increased methylation may ameliorate the disease severity. Aim 2 examines the requirement for DartS, the fly ortholog of PRMT5, in germ cell specification and addresses the specific role played by Sm proteins in this process. Lay summary: Spinal Muscular Atrophy, a common genetic disease, results in very early childhood mortality. Whereas the gene responsible for the disease has been identified, the mechanism that ultimately results in the disease pathology is unknown. In order to develop effective therapies, a molecular understanding of the disease gene product is essential.

描述（由申请人提供）：Sm类小核核糖核蛋白（snRNPs）是剪接体的核心成分，是体内剪接所必需的。此外，Sm蛋白似乎在种系规范中具有更多的祖先作用。该项目的总体目标是了解Sm蛋白被组装成snRNPs的机制，以进行剪接或组装成建立生殖细胞命运所需的特殊核糖核蛋白（RNPs）。在这两种看似不同的通路的连接处是PRMT5复合体。snRNPs的生物发生是高度协调的，需要几个基本的辅助因子。特别是，细胞质snRNP的组装依赖于两种异质复合物，PRMT5复合物和SMN复合物。重要的是，降低SMN蛋白水平的突变与脊髓性肌萎缩症（SMA）的神经退行性疾病有关。患有这种疾病的患者通常在儿童时期就早早死亡。SMA的分子病因尚不清楚，然而，snRNP生物发生的扰动被认为是一个主要的促成因素。对snRNP组装机制的详细了解将有助于阐明该疾病的发病过程。PRMT5复合体负责Sm蛋白的对称二甲基化，并且由于这种活性，被认为与SMN复合体合作介导有效的snRNP组装。然而，目前缺乏对这一过程的体内研究。此外，PRMT5与BLIMP1相关，是小鼠生殖细胞发育所必需的。因此，PRMT5活性的破坏可能会影响两个不同但可分离的途径- snRNP生物发生和生殖细胞发育。为了深入了解这些过程的机理，我们已经开发了利用果蝇作为模型系统以及哺乳动物细胞培养技术的体内方案。目的1探讨snRNP生物发生中Sm蛋白甲基化的需求。由于SMN与甲基化的Sm蛋白具有更高的亲和力，因此这种修饰可能作为SMA表型的修饰因子。具体来说，哺乳动物中Sm蛋白甲基化的缺乏可能会导致SMA的表型。另外，甲基化的增加可能会改善疾病的严重程度。目的2研究了生殖细胞分化过程中对PRMT5的同源基因DartS的需求，并阐述了Sm蛋白在这一过程中所起的特殊作用。摘要：脊髓性肌萎缩症是一种常见的遗传性疾病，导致幼儿死亡。虽然已经确定了导致这种疾病的基因，但最终导致这种疾病病理的机制尚不清楚。为了开发有效的治疗方法，对疾病基因产物的分子理解是必不可少的。