TRANSPLANTATION ANTIGENS--EXPRESSION AND FUNCTION

高度多态经典移植的功能 抗原在移植排斥和抵抗病毒感染中的作用， 长期以来得到了很好的认可。 相比之下， 非多态类大得多的家族的生理效应 I-样MHC分子仍然是未知的。 的结构相似性 这两类多肽表明，非经典MHC 分子可结合自身或非自身肽，并可与T细胞相互作用， 受体。 鉴定蛋白质，从推定的 肽是衍生的，并且认识到它们可能 与之相互作用的细胞代表了重大挑战， 可能导致免疫学新概念的形成。 Qa-2分子是最好的表征非经典 小鼠体内的移植抗原。 它们以两种形式存在：1） 糖磷脂酰肌醇（GPI）连接的膜抗原， 主要在淋巴衍生细胞上表达，并可作为 细胞毒性细胞的靶点以及激活信号的转换器 和2）可溶性Qa-2分子，其被释放到T细胞中。 血清，并已被假设为诱导激活或无反应状态。 从膜到可溶性形式的“转换”在体外是可诱导的， 细胞激活，并涉及Qa-2外显子5的选择性剪接 基因 这项建议的具体目标是： 1. 为了鉴定控制细胞凋亡的Qa-2基因的顺式作用元件， 外显子5的选择性剪接。 2. 为了验证一个假设，各种方案和信号诱导 免疫系统的激活（即，导致选择性剪接 体内Qa-2 mRNA的表达和可溶性Qa-2分子的分泌，以及 为了鉴定参与免疫应答的主要Qa-2表达细胞亚群， 不同的反应。 3. 为了对与分泌的和GPI连接的Qa-2结合的肽进行测序， 分子，以识别由抗原呈递的自身抗原。 两种Qa-2亚型，并对高度 纯化的Qa-2蛋白，以解决3D结构的 非经典MHC抗原。 目标1和2中提议的研究将增进对以下方面的理解： 真核生物可诱导选择性剪接的一般机制 基因，并将提供有关特定条件和细胞的信息， 在免疫应答过程中与优先免疫相关的 产生可溶性或GPI连接的Qa-2产物。 这些数据可以让 我们来定义Qa-2亚型的生理功能。 研究 目标3中的肽将直接检测与这些蛋白质结合的肽， 将评估Qa-2多肽与经典多肽的结构相似性。 移植抗原 结果将用于推断 由Qa-2分子呈递的抗原和所需的参数 MHC分子与蛋白质片段的结合。 从长远来看，拟议中的研究可能有助于设计疫苗， 防止自身免疫和操纵MHC识别过程。

The functions of the highly polymorphic classical transplantation antigens in graft rejection and in defense against viral infections have long been well recognized. In contrast, the properties and the physiological effects of the much larger family of nonpolymorphic class I-like MHC molecules are still unknown. The structural similarities of the two classes of polypeptides suggest that the nonclassical MHC molecules may bind self or nonself peptides and may interact with T-cell receptors. The identification of the proteins from which the putative peptides are derived and the recognition of the effects that they may have on cells with which they interact represent major challenges as they may lead to the formulation of new concepts in immunology. Qa-2 molecules are among the best characterized nonclassical transplantation antigens in the mouse. They exist in two forms: 1) glycophosphatidyl-inositol (GPI) linked membrane antigens which are predominantly expressed on lymphoid derived cells and can serve as targets for cytotoxic cells as well as transducers of activation signals in T cells and 2) soluble Qa-2 molecules which are released into the serum and have been hypothesized to induce activation or anergy states. The "switching" from membrane to soluble form is inducible in vitro by cell activation and involves alternative splicing of exon 5 of the Qa-2 gene. The specific aims of this proposal are: 1. To identify the cis-acting elements of the Qa-2 genes that control alternative splicing of exon 5. 2. To test a hypothesis that various regimens and signals inducing activation of immune system (i.e., lead to the alternative splicing of Qa-2 mRNA and secretion of soluble Qa-2 molecules in vivo, and to identify the major Qa-2 expressing cell subsets participating in different responses. 3. To sequence the peptides bound to the secreted and GPI-linked Qa-2 molecules in order to identify the self-antigens presented by the two Qa-2 isoforms, and to perform crystallography on the highly purified Qa-2 proteins in order to solve a 3D structure of the nonclassical MHC antigen. The studies proposed in Aims 1 and 2 will advance the understanding of the general mechanisms of inducible alternative splicing of eukaryotic genes and will provide information on specific conditions and cell subsets during immune responses that correlate with preferential production of soluble or GPI-linked Qa-2 products. These data may allow us to define the physiological functions of the Qa-2 isoforms. Studies in Aim 3 will examine directly the peptides bound to these proteins and will assess structural similarity of Qa-2 polypeptides to classical transplantation antigens. The results will be used to deduce the antigens presented by Qa-2 molecules and the parameters required for binding of protein fragments by MHC molecules. In the long term the proposed research may help to design vaccines, prevent autoimmunity and manipulate MHC recognition processes.