Role of RNA silencing in B cell development and function

RNA 沉默在 B 细胞发育和功能中的作用

• 批准号: 7367061
• 负责人: KLAUS RAJEWSKY
• 金额: $ 56.79万
• 依托单位: IMMUNE DISEASE INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7367061
• 关键词: 3' Untranslated Regions; Address; Affect; Alleles; Antigen Presentation; Antigens; Apoptosis; Autoimmune Diseases; B-Cell Activation; B-Cell Development; B-Cell Lymphomas; B-Lymphocyte Subsets; B-Lymphocytes; Bioinformatics; Biological; Caenorhabditis elegans; Cell Differentiation process; Cell Lineage; Cell Proliferation; Cells; Chromatin; Chromatin Structure; Complement; DNA Methylation; Dendritic Cells; Dependence; Development; Dicer Enzyme; Double-Stranded RNA; Ectopic Expression; Engineering; Exclusion; Exonuclease; Gene Expression; Gene Rearrangement; Gene Silencing; Gene Targeting; Generations; Genes; Genetic; Green Fluorescent Proteins; Heterochromatin; Histones; Homeostasis; Immune response; Immunity; Immunoglobulin Class Switching; Immunoglobulin Genes; Immunoglobulin Somatic Hypermutation; Immunoglobulin Switch Recombination; Immunoglobulins; Individual; Lower Organism; Lymphocyte; Lymphomagenesis; Malignant - descriptor; Mature B-Lymphocyte; Mediating; Medical; Messenger RNA; MicroRNAs; Molecular; Mus; Mutagenesis; Mutant Strains Mice; Mutation; Nucleotides; Organism; Phase; Play; Process; Property; RNA Interference; Receptor Signaling; Reporter; Resolution; Role; Signal Transduction; Small Interfering RNA; Staging; Stem cells; System; T-Lymphocyte; Testing; Transgenes; Translations; V(D)J Recombination; Work; base; cell type; human DICER1 protein; immune function; in vivo; insight; novel; prevent; promoter; recombinase; research study; self-renewal; small hairpin RNA

DESCRIPTION (provided by applicant): RNA silencing is a novel, major biological mechanism controlling gene expression at the level of messenger RNA and chromatin, triggered by double-stranded RNA. Originally identified in lower organisms, recent results indicate that it also plays a crucial role in mammalian development and cell differentiation. Based on recent evidence that RNA silencing can affect B cell development, and strong indications that its dysregulation contributes to B cell lymphomagenesis, we hypothesize that RNA silencing plays specific roles at various stages of B cell development. The latter is a multi-stage process characterized by a sophisticated ordered interplay of chromosomal loci at which somatic gene rearrangements and mutation occur sequentially, accompanied by phases of cellular proliferation and quiescence and a continuous requirement for survival signals that rescue the cells from apoptosis. Together with the self-renewal capacity of certain mature B cell subsets, some of these features strikingly resemble processes that govern stem cell homeostasis and lineage commitment, known to be prime targets of control by RNA silencing. In the proposed experiments we will introduce targeted mutations into sequential stages of B cell development using Cre/loxP-mediated mutagenesis. Using this approach, we will first investigate the extent to which the RNA silencing machinery is required for proper B cell development and function, including the ability of the cells to execute adaptive immune responses and innate immune functions like antigen presentation to T cells. In the latter context, dendritic cells (DCs) will be included in the analysis. In these experiments we will use a conditional d/cerallele which we have generated, having demonstrated that Dicer is required for RNA silencing in mouse cells. Second, we will investigate whether the RNA silencing machinery is regulated during B cell development, and if so, try to understand the basis of such a control. Finally, we will identify and functionally characterize individual si/miRNAs that are selectively and stage-specifically expressed in B cells and DCs, using bioinformatic target gene predictions as a guide and targeted mutagenesis in mice. Overall, we expect new insights into the control of normal and malignant B cell development and the functional activity of B cells in adaptive and innate immune responses. Given the critical role of B cells in protective immunity, autoimmune diseases and lymphomagenesis, we anticipate this work will have significant medical relevance.

描述（由申请人提供）：RNA沉默是一种新型的主要生物学机制，该机制控制了由双链RNA触发的信使RNA和染色质水平上的基因表达。最初在较低的生物体中鉴定出来，最近的结果表明，它在哺乳动物发育和细胞分化中也起着至关重要的作用。基于最近的证据表明RNA沉默会影响B细胞的发育，并有很强的迹象表明其功能障碍有助于B细胞淋巴细胞增多，我们假设RNA沉默在B细胞发育的各个阶段都在特定的角色中起着特定的作用。后者是一个多阶段过程，其特征是染色体基因座的复杂有序相互作用，在该相互作用，在该染色体基因座中，体细胞基因重排和突变依次发生，伴随着细胞增殖和静脉的相位，以及对凋亡营救细胞的生存信号的连续需求。加上某些成熟B细胞子集的自我更新能力，其中一些特征非常类似于控制干细胞体内稳态和谱系承诺的过程，该过程已知是RNA沉默的主要控制目标。在提出的实验中，我们将使用CRE/LOXP介导的诱变将靶向突变引入B细胞发育的顺序阶段。使用这种方法，我们将首先研究适当的B细胞发育和功能所需的RNA沉默机制，包括细胞执行适应性免疫反应和先天免疫功能（例如抗原对T细胞）的能力。在后一种情况下，树突状细胞（DC）将包括在分析中。在这些实验中，我们将使用有条件的D/Cerallele产生的条件D/cerallele，证明小鼠细胞中的RNA沉默需要DICER。其次，我们将研究在B细胞发育过程中是否调节RNA沉默机制，如果是的，请尝试理解这种对照的基础。最后，我们将使用生物信息学靶基因预测作为指导和靶向诱变，在B细胞和DC中有选择地和阶段特异性表达单个SI/miRNA，并在B细胞和DC中特异性表达。总体而言，我们期望对控制正常和恶性B细胞发育的控制以及B细胞在适应性和先天免疫反应中的功能活性有新的见解。鉴于B细胞在保护性免疫，自身免疫性疾病和淋巴作用中的关键作用，我们预计这项工作将具有很大的医学相关性。