Kinase-mediated assembly of the mRNA entry channel in 40S ribosomes

40S 核糖体中激酶介导的 mRNA 进入通道组装

• 批准号: 9197652
• 负责人: Katrin Karbstein
• 金额: $ 41.65万
• 依托单位: SCRIPPS FLORIDA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9197652
• 关键词: Address; Affect; Beak; Binding; Binding Proteins; Binding Sites; Biochemical; Biochemical Genetics; Biological Assay; Cell Count; Cell physiology; Cells; Colon Carcinoma; Complex; Complex Analysis; Cytoplasm; Data; Defect; Diamond-Blackfan anemia; Disease; Dissection; Dissociation; Electron Microscopy; Ensure; Event; Genetic; Genetic Transcription; Goals; Growth; Incidence; Inherited; Internal Ribosome Entry Site; Laboratories; Lead; Light; Link; Malignant Neoplasms; Masks; Mediating; Messenger RNA; Methods; Modeling; Modification; Patients; Phenotype; Phosphorylation; Phosphotransferases; Protein Analysis; Proteins; Quality Control; RNA; RNA analysis; Recruitment Activity; Ribosomal Proteins; Ribosomal RNA; Ribosomes; Role; Signal Transduction; Site; Solvents; Structure; Syndrome; System; Tail; Testing; Tissues; Translating; Translations; Viral; Work; Yeasts; casein kinase; chromosome 5q loss; cryogenics; experimental study; human disease; insight; leukemia; link protein; novel; osteosarcoma; prevent; public health relevance; rRNA Precursor; reconstitution; tool; tumorigenesis

 DESCRIPTION (provided by applicant): Rapidly dividing cells must produce 2,000 new ribosomes every minute and ensure that they are fully functional. The mRNA entry channel is formed on the beak structure of the small ribosomal subunit, and two out of the three proteins at the beak are associated with DBA, an inherited human disease, caused by haploinsufficiency of ribosomal proteins (r-proteins). As expected from loss of r-proteins, patients have low cell numbers in rapidly dividing tissues, but are also predisposed to cancer, suggesting that these patients fail in producing both the numbers and quality of ribosomes needed. Our recent data indicates that r- proteins at the mRNA entry channel are assembled late during maturation, and that assembly is regulated by release of the assembly factor Ltv1, which requires phosphorylation by the Hrr25 kinase. The overall goal of this project is to dissect the assembly of the mRNA entry channel using structural and biochemical methods. In Aim 1 we will dissect how Ltv1 dissociation affects the structure of pre-40S subunits, and the mRNA binding channel. We will also test how release of Ltv1 is linked to recruitment of the ribosomal protein Asc1, which is required for translation of IRES-containing viral messages. In Aim 2 we will use structural and yeast biochemical methods to define how incorporation of Rps10 at the beak is linked to 18S rRNA processing at the platform, via a network of proteins involved in DBA. These studies will provide insight into assembly events that are very rapid within cells and increase our understanding of the mechanisms by which defects in ribosome maturation can lead to tumorigenesis.

 描述（由适用提供）：迅速分裂的单元必须每分钟产生2,000个新核糖体，并确保它们具有完全功能。 mRNA进入通道是在小核糖体亚基的喙结构上形成的，喙的三种蛋白质中有两种与DBA有关，DBA是一种遗传性的人类疾病，是由核糖体蛋白（R蛋白）的单倍不足引起的。正如R蛋白丧失所预期的那样，患者在快速分裂的组织中的细胞数量较低，但也易于癌症，这表明这些患者无法产生所需的核糖体的数量和质量。我们最近的数据表明，在成熟过程中，mRNA进入通道的R蛋白在晚期组装，并且该组件受释放的组装因子LTV1的释放，这需要HRR25激酶磷酸化。该项目的总体目标是使用结构和生化方法剖析mRNA进入通道的组装。在AIM 1中，我们将剖析LTV1解离如何影响40年代前亚基的结构和mRNA结合通道。我们还将测试LTV1的释放如何与核糖体蛋白ASC1的募集有关，这是含IRES病毒信息的翻译所必需的。在AIM 2中，我们将使用结构和酵母生化方法来定义如何通过涉及DBA的蛋白质网络与平台的18S RRNA处理中的RPS10合并。这些研究将提供对细胞内非常迅速并增加我们的组装事件的见解 了解核糖体成熟中缺陷的机制会导致肿瘤发生。