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The role of N6-methyladenosine RNA modification in programmed and aberrant DNA mutagenesis in B cells

N6-甲基腺苷 RNA 修饰在 B 细胞程序性和异常 DNA 突变中的作用

基本信息

批准号：
10721410
负责人：
Uttiya Basu
金额：
$ 5.31万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-09-09 至 2024-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10721410
关键词：
3-Dimensional Antibodies Antibody Affinity Antigens Automobile Driving B lymphoid malignancy B-Cell Development B-Lymphocytes Biochemistry Biological Assay Biology Bone Marrow CD19 gene Cell Line Cell Lineage Cell physiology Chromatin Chromosomal Instability Chromosome abnormality Code Complex Correlative Study Coupled DNA DNA Double Strand Break DNA Sequence Alteration DNA Single Strand Break DNA Structure Data Defect Disease Enzymes Evaluation Event Frequencies Gene Activation Gene Expression Gene Rearrangement Generations Genes Genetic Recombination Genetic Transcription Genome Genome Stability Genomic Instability Genomics Goals Heavy-Chain Immunoglobulins Hematopoiesis Heterodimerization IGH@ gene cluster IgG1 Immune System Diseases Immunize Immunoglobulin A Immunoglobulin Class Switching Immunoglobulin Genes Immunoglobulin Somatic Hypermutation Immunoglobulin Switch Recombination Immunoglobulins Laboratories Light-Chain Immunoglobulins LoxP-flanked allele Lymphoid Lymphomagenesis Mature B-Lymphocyte Mediating Methylation Microscopy Modification Molecular Multiple Myeloma Mus Mutagenesis Mutation Pathway interactions Peyer&apos s Patches Play Post-Transcriptional RNA Processing Post-Transcriptional Regulation Prevention Process Production Property Proteins RNA RNA Degradation RNA Processing RNA methylation Rag1 Mouse Research Design Resolution Rest Role Single-Stranded DNA Site Somatic Mutation Sterility Structure Structure of germinal center of lymph node System Transcript Untranslated RNA V(D)J Recombination activation-induced cytidine deaminase adaptive immune response cancer cell cellular development ds-DNA epigenomics exosome experimental study genome integrity humoral immunity deficiency in vivo large cell Diffuse non-Hodgkin&apos s lymphoma leukemia/lymphoma mouse genetics mouse model mutant posttranscriptional prevent progenitor recruit single molecule superresolution microscopy transcriptome sequencing

项目摘要

PROJECT SUMMARY Background: VDJ recombination, Class switch recombination (CSR) and somatic hypermutation (SHM) are three B lymphocyte specific processes that mediate antibody gene diversification. VDJ recombination requires the DNA double strand generation by the Recombination activation genes (RAG1 an RAG2) where as CSR and SHM requires the single-strand DNA break activity of the Activation Induced Deaminase (AID) enzyme. Both RAG1/2 and AID activities are coupled with noncoding RNA transcription at sites of DNA break/mutation. The properties of the ncRNAs generated at sites of programmed DNA breaks are poorly characterized in B cells. Recently advances in biology has provided compelling evidence that post-transcriptional and co-transcriptional modification of ncRNAs determine a component of RNA epigenomics and have significant role in driving cellular development and function. In this application, supported by preliminary data generated in our laboratory, we are evaluating the role of RNA modification N6-methyladenosine (m6A) and its associated enzymes METTL3 and METTL14 in B cell development, function and genomic integrity. Objective/Hypothesis: In his proposal, we will determine how RNA methylation m6A on transcripts generated in the IgH locus and the rest of the B cell genome controls programmed DNA recombination, antibody gene diversification and prevents chromosomal instability. Specific Aims: Aim 1: Aim 1: Are m6A modifying enzymes Mettl3 and Mettl14 important for class switch recombination, somatic hypermutation, and/or for preventing genomic stability in B cells? ; AIM 2: What is the mechanism by which m6A modification promotes IgH DNA recombination and/or prevents genomic instability? AIM 3: Do RNA methylation and coupled RNA surveillance pathways have a role in early B cell development and during VDJ recombination? Study Design: Using cell lines and mouse models, we will study mechanism by which RNA m6A methylation plays a role in programmed DNA recombination and protection of B cell genomic integrity. We will evaluate the mechanism of formation of single-strand DNA structures formation and RNA surveillance at sites of AID activity in matured B cells and the mechanism by which inhibitory RNAs are degraded at sites of RAG activity in B cells during VDJ recombination. We will also evaluate the molecular mechanism by which RNA modification promotes programmed DNA rearrangements. We use a combination of mouse genetics, genomics, biochemistry and 3D-STORM microscopy to accomplish the goals of the proposed project. Disease Relevance: B cells are a central component of the adaptive immune response, but also prone to undergo leukemias and lymphomas when antibody gene diversification processes are not well controlled. Proposed studies leads to a better understanding of the mechanism of antibody gene diversification but also educate us how B cell cancer (specially in context of DLBCL and Multiple Myeloma) are prevented.

项目总结