Modulation of B cell tolerance checkpoints by distinct Ras/Erk Pathways

通过不同的 Ras/Erk 通路调节 B 细胞耐受检查点

基本信息

批准号：
8603195
负责人：
Andre Limnander
金额：
$ 6.05万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-01-10 至 2014-09-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8603195
关键词：
1,2-diacylglycerol Address Age Alleles Antibody-Producing Cells Antigens Apoptosis Apoptotic Autoantibodies Autoantigens Autoimmune Diseases Autoimmune Process Autoimmunity B cell repertoire B-Cell Development B-Lymphocytes Biochemical Biochemistry Biological Assay Bone Marrow Cell Line Cells Cessation of life Defect Development Diglycerides Disease EF Hand Motifs Ensure Etiology Event Failure Goals Immature Bone Immune Immunocompetent Immunoglobulin D Immunoglobulin M In Vitro Lead Lymphocyte Lymphopenia Mature B-Lymphocyte Mediating Minor Molecular Mus Mutation Output Pathway interactions Phenotype Phosphorylation Phosphorylation Site Physiological Point Mutation Production Property Protein Biochemistry Protein Isoforms Proteins Relative (related person)Retroviridae Role Serine Signal Pathway Signal Transduction Site Son of Sevenless Proteins Specificity Staging Structure System Systemic Lupus Erythematosus T-Cell Development T-Lymphocyte Testing Tissues cell type in vivo insight lupus-like mouse model mutant novel overexpression public health relevance receptor research study response sensor therapeutic target

项目摘要

DESCRIPTION (provided by applicant): Establishment of a proper B cell repertoire that is immunocompetent but not autoimmune depends on critical developmental checkpoints that edit, silence or delete autoreactive cells. The properties of B cells change as they progress through distinct stages in development, but the signaling mechanisms by which antigen drives the different selection checkpoints are incompletely defined. We have recently described a novel Ca2+- dependent Erk signaling pathway in developing B cells that is pro-apoptotic and mediates B cell negative selection. This pathway requires PKC¿ and RasGRP proteins and loss of this pathway in PKCg-deficient mice results in increased survival of B cells during negative selection and subsequent development of an SLE-like disease with lymphoproliferation and autoantibody production. RasGRP1-deficient mice have a substantial developmental block in T cell development that results in T cell lymphopenia, but as they age they also develop an SLE-like disease with B cell lymphoproliferation and autoantibody production, the etiology of which is not well understood. In addition, a recently identified RasGRP1Anaef allele, which carries a point mutation in the second EF-hand of RasGRP1, also causes an SLE-like disease with distinct effects on T cell development from those observed in the RasGRP1-deficient mice. The first goal of this proposal is to determine whether the SLE-like phenotype in these RasGRP1 mouse models is B cell intrinsic, and if it is due to loss of pro-apoptotic Erk signaling during B cell development. Secondly, because we have identified Serine 332 on RasGRP1 as a putative PKCg target phosphosite that is required for the activation of this novel Ca2+-Erk pathway, I will use in vitro biochemistry experiments to determine the effect of this phosphorylation on the function and specificity of RasGRP1. Finally, I will develop retroviruses encoding mutant S332 RasGRP1 to determine the relevance of this phospho-site in B cell development in vivo. Successful completion of these studies will greatly advance our understanding of Ras/Erk signaling in B cell development in normal and pathological settings. Such insight is essential to define the molecular mechanisms that confer functional specificity to different Ras/Erk pathways, which in turn may pinpoint events that can serve as therapeutic targets while having minor or no consequences on closely related but functionally distinct pathways.

描述（由适用提供）：建立免疫能力但不是自身免疫性的适当的B细胞库，这取决于对编辑，沉默或删除自动反应性细胞的关键发育检查点。 B细胞的特性随着开发的不同阶段而变化，但是抗原驱动不同选择检查点的信号传导机制未完全定义。我们最近描述了一种新型的Ca2+ - 依赖性ERK信号传导途径，该B细胞是促凋亡的B细胞，并介导B细胞阴性选择。该途径需要PKC¿RASGRP蛋白质，并且在PKCG缺陷型小鼠中的该途径丧失会导致在阴性选择期间B细胞的存活增加，并随后伴有淋巴细胞增生和自身抗体产生的SLE样疾病的发展。 RASGRP1缺陷型小鼠在T细胞发育中具有实质性的发育障碍，导致T细胞淋巴增生，但是随着年龄的增长，它们也会发展出具有B细胞淋巴细胞增生和自身抗体产生的SLE样疾病，其病因并不能很好地理解。此外，最近确定的RASGRP1ANAEF等位基因在RASGRP1的第二个EF手中带有点突变，也引起SLE样疾病，对T细胞的发育产生与RASGRP1缺陷小鼠中观察到的疾病的不同作用。该提案的第一个目标是确定这些RASGRP1小鼠模型中的SLE样表型是否是B细胞固有的，并且是否是由于B细胞发育过程中促凋亡的ERK信号传导的丧失所致。其次，由于我们已经将RasGRP1上的丝氨酸332鉴定为一种推定的PKCG靶磷材料，这是激活这一新型Ca2+-erk途径所需的，因此我将使用体外生物化学实验来确定这种磷酸化对RasGRP1功能和特异性的影响。最后，我将开发编码突变体S332 RASGRP1的逆转录病毒，以确定该磷酸位点在体内B细胞发育中的相关性。这些研究的成功完成将大大提高我们对正常和病理环境中B细胞发育中RAS/ERK信号传导的理解。这种洞察力对于定义赋予不同RAS/ERK途径功能特异性的分子机制至关重要，这反过来又可以确定可以用作治疗靶标的事件，而在密切相关但功能上不同的途径上产生较小或没有后果。