Catalytic roles of RNA methyltransferase DIMT1

RNA甲基转移酶DIMT1的催化作用

基本信息

批准号：
10522085
负责人：
BARRY S. COOPERMAN
金额：
$ 60.19万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-07-01 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10522085
关键词：
Ablation Affect Basic Science Biogenesis Biological Biological Assay Cell Line Cell Survival Cells Complex DNA Damage DNA Repair DNA Repair Pathway Data Development Discipline Disease Enzymes Event Fanconi&apos s Anemia Gene Expression Genetic Translation Goals Hematopoiesis Hematopoietic Hematopoietic stem cells Human Human Pathology Impairment In Vitro Internal Ribosome Entry Site Kinetics Knowledge Laboratories Mediating Messenger RNA Methylation Methyltransferase Modification Molecular Pathologic Processes Pathway interactions Peptide Initiation Factors Peptides Physiological Processes Physiology Process Protein Biosynthesis RNA Reaction Regulation Reproducibility Research Ribosomal Frameshifting Ribosomal RNA Ribosomes Role Site Specificity Structure Structure-Activity Relationship System Transfer RNA Translations Variant base cell type cricket paralysis virus insight mRNA Decay novel novel therapeutics polypeptide response scaffold success

项目摘要

Ribosome RNA (rRNA) modifying enzymes are important factors in the assembly and function of ribosomes, dysregulation of which is frequently seen in human hematopoietic disorders. We only have limited knowledge of the catalytic role of rRNA modifying enzymes in the physiological and pathological processes of human hematopoiesis, although a few studies characterized the importance of rRNA modifications in these processes. Preliminary data (CRISPR screening) from my laboratory shows that the ablation of DIMT1 (an 18S rRNA Ni·6A-dimethylation (mi·6A) methyltransferase) significantly impairs cell viability of hematopoietic stem progenitor cells (HSPCs). DIMT1 is highly expressed in HSPC, and its expression decreases in the differentiated lineages. However, the molecular function and detailed mechanism of DIMT1 in human hematopoiesis remain elusive. Even though DIMT1-mediated rRNA modifications are highly conserved, the catalytic activity of DIMT1 is dispensable for proper ribosome biogenesis. However, our recent results suggest that the catalytic activity of DIMT1 is required for the viability of HSPCs, while the catalytic-independent role of DIMT1 is important for 18S rRNA processing and ribosome biogenesis. This implies that DIMT1 may have a necessary role as a scaffold in ribosome biogenesis separate from its catalytic activity. Furthermore, we show that the catalytic role of DIMT1 is indispensable for the expression of genes involved in the Fanconi anemia and other DNA repair pathways. However, whether and how DIMT1-mediated mi·6A installation in 18S rRNA influences cell viability and DNA repair pathways in HSPCs are not known. Here, our goal is to understand the molecular function and the mechanistic details of the catalytic role of DIMT1 in the regulation of cell viability and gene expression in HSPCs. Specifically, Aim 1 will investigate the catalytic role of DIMT1 as an 18S rRNA methyltransferase in HSPC cell viability and protein synthesis. Aim 2 will investigate the molecular mechanism by which DIMT1- mediated m26•6A in 18S rRNA controls translation elongation using an in vitro eukaryotic translation system. Aim 3 will investigate the structure-function relationships and catalytic mechanism of DIMT1. Results from these studies will provide novel insights into how the catalytic role of DIMT1 impacts HSPC cell viability and the expression of genes Fanconi anemia and other DNA repair pathways. The understanding we will gain can be broadly applicable to hematopoietic studies and many other biological disciplines.

核糖体 RNA (rRNA) 修饰酶是核糖体组装和功能的重要因素核糖体的失调在人类造血系统疾病中很常见，我们的研究有限。 rRNA修饰酶在生理和病理过程中的催化作用的知识人类造血过程中，尽管有一些研究描述了 rRNA 修饰在这些过程中的重要性我的实验室的初步数据（CRISPR 筛选）显示 DIMT1（18S）的消融。 rRNA Ni·6A-二甲基化（mi·6A）甲基转移酶）显着损害造血干细胞活力 DIMT1 在 HSPC 中高表达，在分化过程中表达降低。然而，DIMT1 在人类造血中的分子功能和详细机制仍然存在。尽管 DIMT1 介导的 rRNA 修饰高度保守，但其催化活性却难以捉摸。 DIMT1 对于正确的核糖体生物发生是不可或缺的，然而，我们最近的结果表明催化作用。 DIMT1 的活性对于 HSPC 的生存能力是必需的，而 DIMT1 的催化独立作用很重要对于 18S rRNA 加工和核糖体生物合成，这意味着 DIMT1 可能具有作为一个必要的作用。支架在核糖体生物合成中与其催化活性分离，此外，我们还表明其催化作用。 DIMT1 对于参与范可尼贫血和其他 DNA 修复的基因的表达是必不可少的然而，DIMT1 介导的 mi·6A 在 18S rRNA 中的安装是否以及如何影响细胞活力。 HSPC 中的 DNA 修复途径尚不清楚，我们的目标是了解其分子功能和作用。 DIMT1 在细胞活力和基因表达调节中的催化作用的机制细节具体来说，目标 1 将研究 DIMT1 作为 18S rRNA 甲基转移酶的催化作用。 HSPC 细胞活力和蛋白质合成。目标 2 将研究 DIMT1- 的分子机制。 18S rRNA 中介导的 m26•6A 使用体外真核翻译系统控制翻译延伸。 3将研究DIMT1的结构-功能关系和催化机制。研究将为 DIMT1 的催化作用如何影响 HSPC 细胞活力和我们将获得范可尼贫血基因表达和其他 DNA 修复途径的理解。广泛适用于造血研究和许多其他生物学学科。