The role of m6A RNA modification as modulator of dsRNA induced cell-intrinsic innate immune responses in hematopoiesis

m6A RNA 修饰作为 dsRNA 调节剂在造血过程中诱导细胞内在先天免疫反应的作用

基本信息

批准号：
10676211
负责人：
Stephanie Halene
金额：
$ 29.48万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-08-01 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10676211
关键词：
Adult Affect Applications Grants Biological Assay Catalytic Domain Cells Complex Data Defect Double-Stranded RNA Dysmyelopoietic Syndromes Engraftment Failure Fetal Development Fetal Liver Generations Genetic Genetic Transcription Hematopoiesis Hematopoietic Hematopoietic stem cells Immune Response Genes Immune response Immune signaling Immunoprecipitation In Vitro Innate Immune Response Interferons Knock-out Knockout Mice Maps Mediating Messenger RNA Methyltransferase Modeling Modification Nucleotides Pathway interactions Phenotype Play Process Proliferating Proteins RNA RNA metabolism Reader Reporting Resolution Ribonucleases Role Signal Transduction Structure Transcript Transcription Initiation Site Translations Transplantation Up-Regulation Viral Work epitranscriptome functional disability in vivo innate immune pathways knock-down leukemia mouse model novel oligoadenylate pathogen preservation prevent progenitor response sensor single-cell RNA sequencing stem cell function

项目摘要

Project Summary N6-methyladenosine (m6A) is the most abundant RNA modification and plays key roles in RNA metabolism. m6A writers and readers are highly expressed in hematopoietic stem and progenitor cells and dysregulated in myelodysplasia and leukemia. We show that loss of Mettl3 in hematopoietic stem and progenitor cells (HSPCs) resulted in upregulation of an anti-viral innate immune response signature as well as marked proliferation and differentiation defects. In particular, we are the first to report that loss of m6A RNA modification resulted in aberrant dsRNA formation, leading to activation of the MDA5-RIG-I, PKR-eIF2A, and OAS-RNAse L pathways. By differential immunoprecipitation of dsRNAs from Mettl3 KO versus WT cells, we identified a specific subset of long, normally highly m6A modified transcripts with low folding energies, signifying propensity to form extensive RNA secondary structures. Disruption of innate immune signaling via deletion of Mavs or knockdown of Rnasel in part rescued colony formation in vitro and engraftment in competitive transplantation assays in vivo. These data suggest that one of the roles of m6A RNA modifications is to maintain single-strandedness of endogenous RNAs whereby they mark RNAs as “self” and distinguish them from exogenous pathogen-derived dsRNAs. In this SHINE II application we propose to dissect the role of m6A RNA modification in HSPCs specifically as it pertains to its role as suppressor of aberrant innate immune signaling. We will determine the mechanism by which METTL3-mediated m6A RNA modification prevents abnormal dsRNA formation and consequent induction of interferon signaling, we will dissect how m6A loss results in a deleterious immune response that disrupts HSPC function, and we will assess the universality this phenomenon across hematopoietic lineages.

项目概要 N6-甲基腺苷 (m6A) 是最丰富的 RNA 修饰，在 RNA 代谢中发挥关键作用。作者和读者在造血干细胞和祖细胞中高度表达，并在骨髓增生异常和白血病。我们发现造血干细胞和祖细胞 (HSPC) 中 Mettl3 的缺失导致抗病毒先天免疫反应特征以及明显的增殖和分化缺陷。特别是，我们是第一个报告 m6A RNA 修饰缺失导致 dsRNA 形成异常的情况，通过差异导致 MDA5-RIG-I、PKR-eIF2A 和 OAS-RNAse L 途径的激活。通过对来自 Mettl3 KO 细胞和 WT 细胞的 dsRNA 进行免疫沉淀，我们鉴定了一个特定的长链子集，通常为高度 m6A 修饰的转录本具有低折叠能，表明倾向于形成广泛的 RNA 二级结构通过删除 Mavs 或敲低 Rnasel 来破坏先天免疫信号传导，部分得到挽救。体外集落形成和体内竞争性移植测定中的植入这些数据表明。 m6A RNA 修饰的作用之一是维持内源 RNA 的单链性，从而它们将 RNA 标记为“自身”，并将其与外源病原体衍生的 dsRNA 区分开来。在此 SHINE II 应用中，我们建议剖析 m6A RNA 修饰在 HSPC 中的作用，特别是因为它关于其作为异常先天免疫信号抑制剂的作用，我们将通过以下方式确定其机制。 METTL3 介导的 m6A RNA 修饰可防止异常 dsRNA 形成和随后的诱导关于干扰素信号传导，我们将剖析 m6A 缺失如何导致有害的免疫反应，从而破坏 HSPC 功能，我们将评估这种现象在造血谱系中的普遍性。