Differential Signaling by B cell Receptor in Tolerance Induction

B 细胞受体在耐受诱导中的差异信号传导

• 批准号: 8095925
• 负责人: Haochu Huang
• 金额: $ 19.5万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-02-15 至 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8095925
• 关键词: Affect; Animal Model; Antibodies; Antigen Receptors; Antigens; Apoptosis; Autoantigens; Autoimmune Diseases; B cell repertoire; B-Cell Lymphomas; B-Lymphocytes; Binding; Bone Marrow; Candidate Disease Gene; Cell Line; Cell model; Cells; Chronic; Clonal Deletion; DNA; Defect; Event; Gene Targeting; Generations; Genes; Genetic; Genetic Screening; Genomics; Goals; Growth; Immunoglobulins; Knowledge; Libraries; Maintenance; Maps; Membrane; Modeling; Molecular; Mus; Mutation; Nature; Pathway interactions; Patients; Pharmaceutical Preparations; Phosphoric Monoester Hydrolases; Phosphotransferases; Receptor Signaling; Receptors, Antigen, B-Cell; Retroviridae; Role; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Single-Stranded DNA; Specificity; Testing; Transgenic Model; Translating; anergy; antigen binding; autoreactive B cell; crosslink; ds-DNA; genome-wide; in vivo; in vivo Model; inhibiting antibody; mouse genome; novel; receptor; response; small hairpin RNA

DESCRIPTION (provided by applicant): The generation and maintenance of a B cell repertoire that is tolerant to self rely on multiple mechanisms that regulate the differentiation of autoreactive B cells. These mechansims are controlled by the nature and duration of interaction between B cell receptor (BCR) and its antigen. Multivalent membrane-bound self antigens or double-stranded DNA causes immature self-reactive B cells to undergo deletion or receptor editing. Lower valency antigens, such as a soluble antigen or single-stranded DNA, induce anergy. Anergy is characterized by a profound block in proximal antigen receptor signaling resulting in an antigen-unresponsive state. The induction and maintenance of anergy require chronic binding of antigen and signal transduction. Therefore, the strength of BCR signals dictates the fate of developing B cells. However, the molecular events that translate BCR signal strength to fate choice in tolerance induction and underlying signaling pathways are not well understood. We propose to map these signaling pathways genetically using a genome-wide shRNA screen. The rationale of our approach is that induction of a specific form of B cell tolerance will be defective when a component of the underlying signaling pathway is reduced by shRNA knockdown. As in a genetic screen, cells having desired phenotypic changes are first selected after they are transduced with a retroviral shRNA library. Then the shRNAs carried by these cells are amplified and sequenced resulting in the identification of underlying genes. Since it is technically challenging to carry out shRNA screen in vivo, we will first carry out the screen on WEHI-231 B cell lymphoma cell line. Crosslinking of BCR on WEHI-231 cells results in their apoptosis and it has been used as a model of tolerance induction. Our specific aims are i) to identify novel molecules of BCR signaling pathways in WEHI-231 cells; and ii) to establish the roles of identified genes in different in vivo models of B cell tolerance including clonal deletion, receptor editing and anergy. The goal of this project is to identify new components of differential BCR signaling pathways in tolerance induction. Mutations that affect BCR signaling can alter B cell fate decisions and result in autoimmune diseases. Defective tolerance induction has been demonstrated in both animal models and patients of autoimmune diseases. These studies should provide a refined signaling map which will be essential in dissecting the defects of B cell tolerance in autoimmune diseases. Such knowledge should offer new opportunities to develop novel immunomodulatory drugs. PUBLIC HEALTH RELEVANCE: Using a genetic approach, we propose to screen and identify new genes in signaling pathways that are important for establishing or maintaining tolerance in B cells. This refined signaling map will be essential to dissect tolerance defects in autoimmune diseases and to develop novel immunomodulatory drugs.

描述（由申请人提供）：B细胞库的生成和维护自我宽容依赖于调节自动反应性B细胞分化的多种机制。这些机甲受B细胞受体（BCR）及其抗原之间相互作用的性质和持续时间控制。多价膜结合的自抗原或双链DNA会导致不成熟的自反应B细胞进行缺失或受体编辑。较低的价值抗原，例如可溶性抗原或单链DNA，会诱导厌氧。 Anergy的特征是近端抗原受体信号传导的深层阻滞，导致抗原无反发状态。厌食的诱导和维持需要抗原和信号转导的慢性结合。因此，BCR信号的强度决定了发展中B细胞的命运。但是，在耐受性诱导和基础信号通路中将BCR信号强度转化为命运选择的分子事件尚不清楚。 我们建议使用全基因组shRNA筛选在遗传上绘制这些信号通路。我们方法的理由是，当SHRNA敲低降低了基础信号通路的一个成分时，诱导B细胞耐受性的特定形式将是有缺陷的。与遗传筛选一样，具有所需表型变化的细胞在用逆转录病毒SHRNA文库转导后首先选择它们。然后将这些细胞携带的shRNA放大并测序，从而鉴定出潜在的基因。由于在体内执行shrna屏幕在技术上具有挑战性，因此我们将首先在WEHI-231 B细胞淋巴瘤细胞系上进行筛选。 BCR在WEHI-231细胞上的交联导致其凋亡，并已用作耐受性诱导模型。我们的具体目的是i）确定WEHI-231细胞中BCR信号通路的新分子； ii）在不同的B细胞耐受性体内模型中确定已鉴定基因的作用，包括克隆缺失，受体编辑和无反应。该项目的目的是在耐受性诱导中确定差异BCR信号通路的新组件。 影响BCR信号传导的突变会改变B细胞命运的决策并导致自身免疫性疾病。在动物模型和自身免疫性疾病患者中均证明了耐受性诱导。这些研究应提供一个精致的信号图，这对于剖析自身免疫性疾病中B细胞耐受性的缺陷至关重要。这种知识应该为开发新型免疫调节药物提供新的机会。 公共卫生相关性：使用遗传方法，我们建议在信号通路中筛选和识别对于建立或维持B细胞中耐受性至关重要的信号通路中的新基因。该精制信号图对于剖析自身免疫性疾病中的耐受性缺陷和开发新型免疫调节药物至关重要。