MODULATION OF CLASS II ANTIGEN PROCESSING BY HLA DO

主要组织相容性复合体（MHC）II类分子是异二聚体糖蛋白，其结合源自外源蛋白的肽并将其呈递给CD 4 T细胞。 与残留不变链片段相关的MHC II类分子[II类相关不变链肽（CLIP）]通过II类样分子HLA-DM的催化作用负载抗原肽，HLA-DM也起II类分子伴侣和肽编辑器的作用。 HLA-DO（DO）是另一种在B细胞、树突细胞和胸腺上皮中表达的II类分子，在转运至MHC II类区室（MIIC）期间和之后与DM物理结合。 MIIC中的DM-DO缔合表明两种分子的功能是相连的。 DO显示阻断DM功能，因为纯化的DM-DO复合物不能在体外催化CLIP交换肽。 DO在II类+ DO-细胞系中的表达引起II类-CLIP复合物在细胞表面的积累，表明DO在体内阻断DM功能。DM功能的抑制通过降低细胞表面II类肽复合物的水平导致II类加工途径的下调。所观察到的DO介导的DM功能抑制的生物化学、功能和免疫学后果仍然未知。此外，尚不清楚DO是否具有其他功能。本申请中提出的研究旨在定义DO表达的生物学后果，并确定在II类加工途径中是否存在DO的替代功能。 这将通过三种不同的生物化学方法来实现。 首先，将产生在存在和不存在HLA-DO的情况下表达II类抗原加工途径的已知组分的真正抗原呈递细胞系，并使用生物化学方法进行分析。 第二，我们产生并检测了在没有DM的情况下可以从ER转运但保留其与DM缔合的能力的HLA-DO分子，以分析DO结构和功能。第三，将确定B细胞受体的内化或B细胞的活化是否导致Do从DM释放，从而活化DM。 拟议的工作应该回答有关DO的作用的重要问题，第二类加工途径，并在肿瘤监测和自身免疫等领域的相关性。

Major histocompatibility complex (MHC) class II molecules are heterodimeric glycoproteins that bind peptides derived from exogenous proteins and present them to CD4 T cells. MHC class II molecules associated with residual invariant chain fragments [class II-associated invariant chain peptides (CLIP)] are loaded with antigenic peptides by the catalytic action of the class II- like molecule, HLA-DM that also functions as a class II chaperone and peptide editor. HLA-DO (DO), yet another class II-like molecule that is expressed in B cells, dendritic cells and thymic epithelium, physically associates with DM both during and after transport to the MHC class II compartments (MIIC). The DM-DO association in the MIIC suggested that the functions of both molecules are linked. DO was shown to block DM function as purified DM-DO complexes could not catalyze CLIP exchange for peptide in vitro. Expression of DO in class II+ DO- cell lines caused the accumulation of class II-CLIP complexes at the cell surface, suggesting that DO blocked DM function in vivo. The inhibition of DM function results in down modulation of the class II processing pathway by decreasing the level of class II-peptide complexes on the cell surface. The biochemical, functional and immunological consequences of the observed DO-mediated inhibition of DM function remain unknown. Additionally, it is not known if DO has additional functions. The studies proposed in this application are designed to define the biological consequences of DO expression and to determine if there are alternative functions for DO in the class II processing pathway. This will be accomplished by three different biochemical approaches. First, bonafide antigen presenting cell lines that express the known components of the class II antigen processing pathway in the presence and absence of HLA-DO will be generated and analyzed using a biochemical approach. Second, HLA-DO molecules that can transport from the ER in the absence of DM but that retain their ability to associate with DM we be generated and examined to analyze DO structure and function. Third, it will be determined if internalization of the B cell receptor or if activation of B cells results in the release of Do from DM, thereby activating DM. The proposed work should answer important questions about the role of DO the class II processing pathway and has relevance in such areas as tumor surveillance and autoimmunity.