Characterization of the MHC Class II Peptide Loading Complex

描述(申请人提供)：B细胞和CD4T细胞之间的相互作用是由多肽-MHC II类复合体介导的，并支持体液免疫反应的充分发展。B细胞是抗原特异性的抗原提呈细胞，在B细胞受体(BCR)介导的同源抗原的结合和内化之后，免疫相关的抗原处理发生。本实验室已经证实，BCR介导的抗原加工发生在抗原(Ag)-BCR泛素化之后，并导致表达具有独特功能和生化特性的衍生多肽-II类复合体(称为I型复合体)。推动这个项目的基本假设是，Ag-BCR复合体的加工发生在MHC II类多肽负载复合体(PLC)中，该复合体位于MHC II类富含抗原的处理舱中。为了验证这一假设，我们将扩展我们的新的初步数据，并采用生化方法进一步定义B细胞中处理通过BCR介导的或液体相内吞作用(目标1)内化的抗原的II类PLC的分子组成。我们还将利用“FRET显微镜”方法来研究完整B细胞中第二类PLC形成的动力学(目标2)。这个项目的总体目标是 建议更好地了解BCR介导的抗原处理后第二类多肽负载的分子机制，并确定第二类多肽负载是否发生在含有特定抗原肽来源的PLC中(即，Ag-BCR复合体)。 公共卫生相关性：抗原特异性B细胞和CD4辅助T细胞之间的相互作用受到B细胞表达的多肽-MHC II类复合体的限制。该项目的目标是确定抗原特异性B细胞产生这些关键的多肽-II类复合体的分子机制。这些信息将有助于新疫苗的设计 增强这些复合体的产生或治疗方案，以抑制自身免疫情况下的这一机制。

DESCRIPTION (provided by applicant): Interactions between B cells and CD4 T cells are mediated by peptide-MHC class II complexes and support full development of a humoral immune response. B cells are antigen-specific antigen presenting cells, where immunologically relevant antigen processing occurs subsequent to B cell receptor (BCR)-mediated binding and internalization of cognate antigen. This laboratory has established that BCR-mediated antigen processing occurs subsequent to antigen (Ag)-BCR ubiquitination and results in expression of derivative peptide-class II complexes (termed "Type I" complexes) with unique functional and biochemical properties. The underlying hypothesis driving this project is that processing of Ag-BCR complexes occurs within an MHC class II peptide-loading complex (PLC) located in MHC class II enriched antigen processing compartments. To test this hypothesis, we will extend our new preliminary data and take a biochemical approach to further define the molecular composition of the class II PLC in B cells processing antigen internalized either via BCR-mediated or fluid-phase endocytosis (Aim 1). We will also utilize a "FRET microscopy" approach to study the dynamics of class II PLC formation in intact B cells (Aim 2). The overall goal of this proposal is to gain a better understanding of the molecular mechanism of class II peptide loading subsequent to BCR-mediated antigen processing, and to determine if class II peptide loading occurs within a PLC containing a dedicated source of antigenic peptide (i.e., Ag-BCR complexes). PUBLIC HEALTH RELEVANCE: Interactions between antigen specific B cells and CD4 helper T cells are restricted by B cell expressed peptide-MHC class II complexes. The goal of this project is to define the molecular mechanism by which antigen specific B cells generate these critical peptide-class II complexes. This information will be helpful in the design of new vaccines that enhance the production of these complexes or therapeutic protocols to dampen this mechanism in cases of autoimmunity.