Characterization of the MHC Class II Peptide Loading Complex

描述（由申请方提供）：B细胞和CD 4 T细胞之间的相互作用由肽-MHC II类复合物介导，并支持体液免疫应答的全面发展。B细胞是抗原特异性抗原呈递细胞，其中免疫相关抗原加工发生在B细胞受体（BCR）介导的同源抗原的结合和内化之后。该实验室已经确定，BCR介导的抗原加工发生在抗原（Ag）-BCR泛素化之后，并导致具有独特功能和生物化学性质的衍生肽II类复合物（称为“I型”复合物）的表达。驱动该项目的基本假设是，Ag-BCR复合物的加工发生在位于MHC II类富集抗原加工区室中的MHC II类肽加载复合物（PLC）内。为了验证这一假设，我们将扩展我们的新的初步数据，并采取生物化学方法，以进一步确定在B细胞处理通过BCR介导的或液相内吞作用内化的抗原中II类PLC的分子组成（目的1）。我们还将利用“FRET显微镜”方法来研究完整B细胞中II类PLC形成的动力学（目的2）。这个项目的总体目标是 该建议是为了更好地理解BCR介导的抗原加工后II类肽加载的分子机制，并确定II类肽加载是否发生在含有抗原肽的专用来源的PLC内（即，Ag-BCR复合物）。

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).