Coordination of SR Protein Phosphorylation and RNA Splicing

SR 蛋白磷酸化和 RNA 剪接的协调

• 批准号: 7913861
• 负责人: JOSEPH ADAMS
• 金额: $ 16.54万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-09 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7913861
• 关键词: 3' Splice Site; Address; Affinity; Alternative Splicing; Arginine; Ataxia; Biological Assay; Biological Process; C-terminal; Cell Nucleus; Cells; Cellular Assay; Chemicals; Cisplatin; Communities; Complex; Cytoplasm; Dipeptides; Disease; Docking; Down-Regulation; Drug Delivery Systems; Engineering; Enzymes; Event; Family; Funding; Genes; Goals; Health; Hereditary Disease; Human; Human Genome; Kinetics; Lead; Link; Lower Organism; Malignant Neoplasms; Mental disorders; Messenger RNA; Methods; Modification; Molecular Conformation; Muscular Dystrophies; Mutation; N-terminal; Names; Neurodegenerative Disorders; Neurofibromatoses; Organelles; Pancreatic carcinoma; Parkinsonian Disorders; Pathway interactions; Pattern; Phosphorylation; Phosphorylation Site; Phosphotransferases; Protein Binding; Protein Conformation; Protein Family; Protein Kinase; Proteins; RNA; RNA Processing; RNA Recognition Motif; RNA Splicing; Reaction; Refractory; Retinoblastoma; Role; Serine; Site; Specificity; Spliced Genes; Spliceosome Assembly Pathway; Spliceosomes; Structure; Testicular Neoplasms; Testing; chemotherapeutic agent; chemotherapy; human disease; interest; mRNA Precursor; macromolecular assembly; novel; protein function; protein structure; prototype; public health relevance; treatment strategy; tumor

DESCRIPTION (provided by applicant): Comparisons of the genomes from humans and lower organisms reveal that the complexity in humans is achieved not by a dramatic increase in the number of genes but by alternative splicing events that stitch together different portions of genes to generate diverse proteins. Correct splicing allows normal healthy function, however, incorrect splicing is linked to many human diseases. For example, muscular dystrophy, ataxias, parkinsonism, neurofibromatosis, psychiatric disorders and cancer have their origins in splicing errors. Splicing reactions are catalyzed by a large macromolecular machine known as the spliceosome. Composed of both RNA and protein, the spliceosome can accurately select the proper splice sites from a considerably large precursor mRNA in healthy cells. The assembly of the spliceosome, the identification of the correct 5' and 3' splice sites and the chemical splicing reaction itself is regulated by a large class of splicing factors known as SR proteins. SR proteins contain one or two RNA recognition motifs and a long C-terminal domain rich in numerous arginine-serine dipeptide repeats. The phosphorylation of the RS domain serves many RNA processing functions including splice-site selection, import of SR proteins into the nucleus and export of mature mRNA to the cytoplasm. This project will investigate how two principal families of splicing enzymes uniquely impact SR protein function through regiospecific, multi-site phosphorylation of the RS domains. Using engineered footprinting methods, the directionality of the splicing enzymes will be defined and shown to control which serines in the RS domain are modified. The effects of these selective phosphorylation reactions on SR protein structure and interaction/function within the spliceosome will then be evaluated using kinetic, structural, splicing and cellular assays. The goal is to identify how splicing kinases recognize and phosphorylate specific regions of the RS domains and determine how these chemical modifications impact splicing componentry. PUBLIC HEALTH RELEVANCE: Health Relevance Statement Comparisons of the genomes from humans and lower organisms reveal that the complexity in humans is achieved not by a dramatic increase in the number genes but by splicing events that stitch together different portions of genes to generate diverse proteins. Correct splicing allows normal healthy function, however, incorrect splicing is linked to many human neurodegenerative diseases and cancer. We are investigating how the newly identified drug targets for diverse diseases known as splicing enzymes (named SR-kinases) regulate important splicing factors (a specific family of proteins known as SR proteins) which cooperate in the control of alternative splicing reactions important in both health and disease.

描述（由申请人提供）：对人类和低等生物基因组的比较表明，人类的复杂性不是通过基因数量的急剧增加实现的，而是通过将基因的不同部分拼接在一起以产生多种蛋白质的选择性剪接事件实现的。正确的剪接允许正常的健康功能，然而，错误的剪接与许多人类疾病有关。例如，肌肉萎缩症、共济失调、帕金森氏症、神经纤维瘤病、精神疾病和癌症都起源于剪接错误。剪接反应是由一个被称为剪接体的大分子机器催化的。剪接体由RNA和蛋白质组成，可以在健康细胞中从相当大的前体mRNA中准确地选择合适的剪接位点。剪接体的组装、正确的5‘和3’剪接位点的识别以及化学剪接反应本身都是由一类被称为SR蛋白的剪接因子调节的。SR蛋白含有一个或两个RNA识别基序和一个富含大量精氨酸-丝氨酸二肽重复序列的长c端结构域。RS结构域的磷酸化具有多种RNA加工功能，包括剪接位点选择、SR蛋白输入细胞核和成熟mRNA输出到细胞质。该项目将研究两个主要的剪接酶家族如何通过区域特异性的RS结构域的多位点磷酸化来独特地影响SR蛋白的功能。使用工程足迹方法，剪接酶的方向性将被定义和显示，以控制哪些丝氨酸在RS结构域被修饰。这些选择性磷酸化反应对SR蛋白结构和剪接体内相互作用/功能的影响将随后通过动力学、结构、剪接和细胞分析进行评估。目的是确定剪接激酶如何识别和磷酸化RS结构域的特定区域，并确定这些化学修饰如何影响剪接成分。对人类和低等生物基因组的比较表明，人类的复杂性不是通过基因数量的急剧增加实现的，而是通过将基因的不同部分拼接在一起以产生多种蛋白质的剪接事件实现的。正确的剪接允许正常的健康功能，然而，不正确的剪接与许多人类神经退行性疾病和癌症有关。我们正在研究新发现的用于多种疾病的剪接酶（称为SR激酶）的药物靶点如何调节重要的剪接因子（称为SR蛋白的特定蛋白质家族），这些剪接因子在控制对健康和疾病都重要的替代剪接反应中相互合作。