Mechanism and Regulation of eIF6 in Translation

eIF6翻译机制及调控

• 批准号: 10444189
• 负责人: Sofia Origanti
• 金额: $ 31.12万
• 依托单位: SAINT LOUIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-07 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10444189
• 关键词: Address; Affect; Allosteric Regulation; Aminoacyl-tRNA hydrolase; Binding; Biochemical; Biological Assay; Biological Models; Biophysics; Cell Cycle; Cell Nucleolus; Cell Survival; Cells; Cellular Assay; Childhood Leukemia; Colon Carcinoma; Complex; Data; Deuterium; Disease; Disease Progression; Dissociation; Ensure; Eukaryotic Initiation Factors; Event; GTP Binding; Generations; Goals; Growth; Growth and Development function; Guanosine Triphosphate Phosphohydrolases; Human; Hydrogen; Hydrolysis; Impairment; Inherited; Link; Malignant Neoplasms; Mass Spectrum Analysis; Mediating; Messenger RNA; Metabolic Diseases; Methodology; Mitosis; Modeling; Molecular; Mutation; Pathogenesis; Pathologic; Patients; Peptide Initiation Factors; Phenotype; Phosphorylation; Physiological; Process; Prognosis; Proliferating; Protein Biosynthesis; Protein Deregulation; Quality Control; RPL10 gene; Regulation; Ribonucleoproteins; Ribosomal RNA; Ribosomes; Role; Shwachman-Diamond syndrome; Site; Tail; Testing; Therapeutic; Time; Tissue Microarray; Trans-Activators; Translation Initiation; Translations; Tumor-Derived; Up-Regulation; Work; aurora B kinase; base; cancer cell; cancer subtypes; cancer type; experimental study; in vivo; insight; leukemia; malignant breast neoplasm; mouse model; mutant; new therapeutic target; novel; overexpression; release factor; single molecule; therapeutic development; therapeutic target; triple-negative invasive breast carcinoma; tumorigenesis

PROJECT SUMMARY Ribosomes are large ribonucleoprotein complexes that are integral to translational control. The assembly and maturation of ribosomal subunits involves a multitude of trans-acting factors that render the subunits translationally competent. Maturation of the 60S ribosomal subunit is accomplished by the release of eukaryotic initiation factor-6 (eIF6) with the help of the maturation factors-SBDS and EFL1-GTPase. eIF6 sterically hinders association of the 60S and 40S subunits and therefore, its release from 60S is critical to permit interactions between 60S and the mRNA-bound 40S subunits. In addition, eIF6 is essential for rRNA processing in the nucleolus and is associated with the translationally stalled 60S-ribosome quality control complex. Given its essential roles, the spatial and temporal aspects of eIF6 activities and its release from 60S must be tightly regulated to ensure successful initiation of translation. Impaired release of eIF6 is the defining hallmark of certain ribosomopathies: Shwachman-Diamond syndrome and RPL10 mutations-driven pediatric leukemias. eIF6 levels are also deregulated in several cancers and its enhanced expression is associated with a poor prognosis. Remarkably, restricting eIF6 levels inhibits growth of certain cancers without affecting normal growth. Therefore, targeting eIF6 and its release from 60S has been proposed to be a desirable therapeutic strategy for cancers and ribosomopathies. However, we are yet to understand the role of eIF6 in modulating the distinct steps of 60S assembly and maturation. Also, the molecular mechanisms that regulate eIF6 interactions with the 60S are not completely understood. Towards understanding these mechanisms, our recent work has identified key residues in critical interfaces of eIF6 that modulate its interaction with 60S, and our preliminary data provide direct evidence that the disruption of this interaction is detrimental to cancer cell viability. We have also identified novel sites of regulation in the C-terminus of eIF6 and have uncovered its importance for controlling translational rates. Building on these discoveries, in the current proposal we aim to define the mechanistic steps that promote the release of eIF6 by SBDS, and EFL1 from distinct functional states of 60S and to uncover the role of GTPase activity of EFL1 using a rigorous set of biochemical, biophysical, and single molecule approaches. In addition, we aim to uncover the mechanism of allosteric regulation by the C-terminus of eIF6 and elucidate the functional and phenotypic effects of disease-specific mutations of eIF6 using cellular and in vivo approaches. These studies will provide crucial mechanistic insight into 60S dynamics and will enable the development of therapeutics focused on eIF6 and its regulators.

项目摘要 核糖体是大型核糖核蛋白络合物，是翻译控制不可或缺的。组装和 核糖体亚基的成熟涉及多种导致亚基的反式作用因子 翻译能力。 60年代核糖体亚基的成熟是通过释放真核生物来完成的 启动因子-6（EIF6）借助成熟因子-SBD和EFL1-GTPase。 EIF6在空间上阻碍 60年代和40年代亚基的关联，因此，它从60年代释放对于允许互动至关重要 在60年代和连接mRNA的40S亚基之间。此外，EIF6对于在RRNA处理中是必不可少的 核仁与翻译停滞的60s-核糖体质量控制复合物有关。鉴于它 基本角色，EIF6活动的空间和时间方面及其从60年代释放必须紧密 受监管以确保成功开始翻译。 EIF6的释放受损是某些的定义标志 核糖体病：Shwachman-Diamond综合征和RPL10驱动的小儿白血病。 EIF6级别 在几种癌症中也被放松管制，其增强表达与预后不良有关。 值得注意的是，限制EIF6水平会抑制某些癌症的生长而不会影响正常生长。所以， 针对EIF6及其从60年代释放的目标是癌症的理想治疗策略 和核糖体病。但是，我们尚未了解EIF6在调节60年代不同步骤中的作用 组装和成熟。同样，调节EIF6相互作用与60s的分子机制不是 完全理解。为了理解这些机制，我们最近的工作确定了关键残留物 在EIF6的关键接口中，调节其与60s的相互作用，我们的初步数据可直接 证据表明这种相互作用的破坏对癌细胞的生存力有害。我们还确定了小说 EIF6的C端监管部位，并发现了其控制翻译率的重要性。 在这些发现的基础上，在当前的建议中，我们旨在定义促进该的机械步骤 SBD释放EIF6，而EFL1从60s的不同功能状态中释放，并揭示GTPase的作用 使用一组严格的生化，生物物理和单分子方法的EFL1活性。此外， 我们旨在揭示EIF6的C端变构调节的机制，并阐明功能 使用细胞和体内方法，EIF6的疾病特异性突变的表型效应。这些研究 将为60年代的动态提供关键的机械洞察力，并能够开发治疗剂 专注于EIF6及其监管机构。