Structural studies of RNA processing and ubiquitin-like protein modification

RNA加工和类泛素蛋白修饰的结构研究

• 批准号: 9294090
• 负责人: CHRISTOPHER D. LIMA
• 金额: $ 43.98万
• 依托单位: SLOAN-KETTERING INST CAN RESEARCH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-10 至 2021-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9294090
• 关键词: Address; Biochemical Genetics; Biology; Cell Cycle Progression; Cell Cycle Proteins; Cell Cycle Regulation; Cell physiology; Cells; Chemicals; Chromosome Segregation; Complex; Coupled; Cytokinesis; Defect; Disease; Enzymes; Equilibrium; Eukaryota; Exoribonucleases; Genes; Genetic Transcription; Genetic study; Health; Homeostasis; Human; In Vitro; Individual; Inflammation; Life; M cell; Malignant Neoplasms; Metabolism; Mission; Modification; Molecular; Monitor; Nerve Degeneration; Neurodegenerative Disorders; Nuclear; Pathway interactions; Peptide Hydrolases; Play; Post-Translational Protein Processing; Process; Protein Complex Subunit; Proteins; RNA; RNA Decay; RNA Degradation; RNA Processing; Recruitment Activity; Role; Saccharomycetales; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Structure; Transcriptional Regulation; Ubiquitin; Ubiquitin Like Proteins; United States National Institutes of Health; Yeasts; enzyme mechanism; exosome; human disease; in vivo; nucleic acid metabolism; nucleocytoplasmic transport; protein degradation; public health relevance; receptor; reconstitution; ubiquitin-protein ligase

 DESCRIPTION (provided by applicant): Ubiquitin-like proteins. Signal transduction pathways rely on reversible chemical modifications to relay information within and across cells. Covalent modification of protein substrates by ubiquitin and the ubiquitin-like proteins such as SUMO (small ubiquitin-like modifier) contribute to pathways that regulate cellular functions including nuclear transport, cytokinesis, chromosome segregation, G2-M cell cycle progression and transcriptional regulation among many others. Post-translational modification by ubiquitin (Ub) and ubiquitin-like (Ubl) proteins requires the sequential action of E1 activating enzymes, E2 conjugating enzymes and E3 ligases while Ub/Ubl processing and deconjugation is catalyzed by Ub/Ubl-specific proteases. Ubiquitin and SUMO conjugation play an integral role in eukaryotic nuclear metabolism and cell cycle control and our studies are of direct relevance to human health, cancer, and the mission of the NIH. This proposal seeks to address the functional significance for components of the ubiquitin and SUMO conjugation pathways through structural, biochemical and genetic studies that will establish the basis for Ub/Ubl 1) activation, 2) conjugation by E2 and E3 enzymes, 3) and signal transduction through characterization receptors that recognize Ub/Ubl-conjugated substrates. The enzymes, mechanisms and factors that constitute ubiquitin and SUMO protein conjugation pathways are conserved so our studies are broadly relevant to other Ub/Ubl-related pathways. RNA decay. Shifting the balance between RNA transcription and degradation regulates RNA lifetime, quality and abundance. Two principle RNA decay pathways exist in eukaryotes, one catalyzes degradation 5' to 3' while the other degrades RNA in the 3' to 5' direction. The 3' to 5' decay pathway requires the activities of the RNA exosome, a large multi-subunit protein complex that contains a non-catalytic core of nine subunits and two additional subunits that catalyze processive and distributive 3' to 5' RNA exoribonuclease activities. In budding yeast, ten of the eleven genes are essential for life, suggesting the importance of the RNA exosome and its activities in cellular function. Recent efforts illuminated fundamental aspects of eukaryotic exosome structure and function, however many questions remain with respect to the collective activities for exosome subunits in RNA processing and decay and how they are coupled to factors the recruit and modify the RNA substrates. RNA decay pathways play an integral role in eukaryotic nucleic acid metabolism, so our studies are of direct relevance to human health and the mission of the NIH because misregulation of RNA processing and decay is associated with diseases such as cancer, inflammation and neurodegeneration. This proposal will address central issues of human and yeast RNA exosome biology by characterizing individual exosome subunits, by reconstituting multi-subunit RNA exosomes and by analyzing the activities of these complexes in biochemical, genetic and structural studies that will establish functions for the exosome during RNA processing and degradation in vitro and in vivo.

 性状（由申请人提供）：泛素样蛋白。信号转导途径依赖于可逆的化学修饰来在细胞内和细胞间传递信息。泛素和泛素样蛋白如SUMO（小泛素样修饰物）对蛋白质底物的共价修饰有助于调节细胞功能的途径，包括核转运、胞质分裂、染色体分离、G2-M细胞周期进展和转录调控等。泛素（ubiquitin，Ub）和类泛素（ubiquitin-like，Ubl）蛋白的翻译后修饰需要E1激活酶、E2结合酶和E3连接酶的顺序作用，而Ub/Ubl的加工和解结合则由Ub/Ubl特异性蛋白酶催化。泛素和SUMO结合在真核细胞核代谢和细胞周期控制中起着不可或缺的作用，我们的研究与人类健康、癌症和NIH的使命直接相关。该提案试图通过结构、生物化学和遗传学研究来解决泛素和SUMO缀合途径组分的功能意义，这些研究将建立Ub/Ubl 1）活化的基础， 2)通过E2和E3酶的缀合，3）和通过识别Ub/Ubl缀合的底物的表征受体的信号转导。构成泛素和SUMO蛋白缀合途径的酶、机制和因子是保守的，因此我们的研究与其他Ub/Ubl相关途径广泛相关。RNA衰变。改变RNA转录和降解之间的平衡调节RNA的寿命，质量和丰度。真核生物中存在两种主要的RNA降解途径，一种催化5'至3'降解，而另一种在3'至5'方向降解RNA。3'至5'衰变途径需要RNA外泌体的活性，RNA外泌体是一种大型多亚基蛋白质复合物，其含有9个亚基的非催化核心和催化进行性和分配性3'至5' RNA核糖核酸外切酶活性的两个另外的亚基。在芽殖酵母中，11个基因中有10个是生命所必需的，这表明RNA外泌体及其在细胞内的活性的重要性。 功能最近的努力阐明了真核外泌体结构和功能的基本方面，然而关于外泌体亚基在RNA加工和衰变中的集体活动以及它们如何与募集和修饰RNA底物的因子偶联仍然存在许多问题。RNA降解途径在真核核酸代谢中起着不可或缺的作用，因此我们的研究与人类健康和NIH的使命直接相关，因为RNA加工和降解的失调与癌症，炎症和神经退行性疾病等疾病有关。该提案将通过表征单个外泌体亚基、通过重构多亚基RNA外泌体以及通过在生物化学、遗传学和结构研究中分析这些复合物的活性来解决人类和酵母RNA外泌体生物学的中心问题，这些研究将建立外泌体在生物化学、遗传学和结构研究中的功能。 RNA在体外和体内的加工和降解。