PEPTIDE BINDING AND EXPRESSION OF MHC CLASS I MOLECULES

目前主要的抗原呈递的共识是， 组织相容性复合体（MHC）I类分子的一个重要特征是，这些分子 结合来自外源或自身蛋白质的肽， 呈递细胞 然后，这些肽与MHC联合呈递， I类分子与抗原特异性、MHC I类分子限制性T CD8+细胞毒性T淋巴细胞（CTL）上的细胞受体;这种相互作用 最终应导致呈递细胞或靶细胞的细胞溶解， CTL. MHC I类分子的晶体学分析显示， 其中肽被提议结合和呈递;此外， 这些研究提示了MHC I类分子沟中的潜在残基 能够与抗原肽相互作用的分子。 进行的研究 汤森等人的研究表明，抗原肽在 第一类分子的折叠和组装;此外，这些研究 这表明，这一过程导致出口从ER到细胞表面。 然而，Ploegh和同事最近的研究已经产生了一个 关于机制（及其生理相关性）的争议 通过该过程可以发生，因为他们能够证明I类 分子能够在细胞表面进行肽诱导的组装。 我们 先前已经证明，鼠I类分子Ld是 转运到细胞表面的速度更慢，与β 2-m的结合更少 强，并显示较低的表面表达比大多数其他MHC I类 汉森和他的同事表明，表面表达的Ld 当细胞与LD限制性抗原孵育时， 缩氨酸 最近，我们鉴定了一种单克隆抗体， 的Ldalt，其可以代表Ld的未组装前体。 因此，Ld 代表了研究抗原呈递作用的理想模型系统 在MHC I类分子的表面表达中。 的第一个目标 建议是确定Ld的哪些氨基酸残基负责 通过分析Ld样蛋白的表达， 分子，并通过创建和研究嵌合和 突变Ld分子。 第二，为解决上述争议，我们建议 评估上述Ldalt和β 2-m在组装中的作用 过程，并检查动力学，如果体内肽诱导 过程 第三，我们建议分析Ld，Ld样， 具有抗原肽和取代肽的突变Ld分子， 几种不同的测定，包括体外肽结合测定。 这些分析应确定哪些Ld残基参与肽 有约束力，并应导致更好地了解"地形"的 肽-I类分子相互作用。 总体而言， 这些研究将有助于阐明抗原肽所起的作用 在I类抗原的表面表达和随后的抗原呈递中 分子以及I类分子上的残基， 这些过程。

The current consensus view of antigen presentation by major histocompatibility complex (MHC) class I molecules is that these molecules bind peptides derived from foreign or self-proteins synthesized within the presenting cell. The peptides are then presented in association with MHC class I molecules to antigen-specific, MHC class I molecule-restricted T cell receptor on CD8+ cytotoxic T lymphocytes (CTL); this interaction ultimately should result in cytolysis of the presenting or target cell by CTL. Crystallographic analyses of MHC class I molecules revealed a groove into which peptides are proposed to be bound and presented; furthermore, these studies suggested potential residues in the groove of MHC class I molecules that could interact with antigenic peptides. Studies performed by Townsend et al. have implicated a role for antigenic peptide in the folding and assembly of class I molecules; furthermore, these studies suggested that this process led to egress from the ER to the cell surface. However, recent studies by Ploegh and coworkers have generated a controversy concerning the mechanism(s) (and its physiological relevance) by which that process can occur, since they were able to show that class I molecules are capable of peptide-induced assembly at the cell surface. We have previously demonstrated that the murine class I molecule, Ld, is transported to the cell surface more slowly, associates with beta2-m less strongly and displays lower surface expression than most other MHC class I molecules; Hansen and colleagues showed that the surface expression of Ld could be increased upon incubation of cells with Ld-restricted antigenic peptides. Recently, we characterized a mAb that recognizes an altered form of Ldalt that may represent an unassembled precursor of Ld. Thus, Ld represents an ideal model system to study the role of antigen presentation in surface expression of MHC class I molecules. The first objective of the proposal is to determine which amino acid residues of Ld are responsible for its expression phenotype by analyzing the expression of Ld-like molecules, and by creating and studying the expression of chimeric and mutant Ld molecules. Second, to resolve the above controversy, we propose to assess the role of aforementioned Ldalt and beta2-m in the assembly process, and to examine the kinetics if the in vivo peptide-induction process. Third, we propose to analyze the interaction of Ld, Ld-like, and mutant Ld molecules with antigenic peptides and substituted peptides in several different assays including an in vitro peptide binding assay. These analyses should determine which Ld residues are involved in peptide binding and should lead to be a better understanding of the "topography" of the peptide-class I molecule interaction. Collectively, completion of these studies should help elucidate the role that antigenic peptides play in surface expression of and subsequent antigen presentation by class I molecules and as well as the residues on class I molecules involved in these processes.