ANTIGEN PRESENTING CELLS OF THE SKIN

该项目将通过以下方式研究皮肤抗原提呈细胞 确定CD1a的功能，CD1a是一种非多态分子，由 朗格汉斯细胞(LC)。CD1a与I类专业的相似之处 组织相容性(MHC)分子提示CD1a可结合肽 抗原。与I类MHC分子一样，通过T细胞介导细胞毒作用 抗原的受体(TCR)，但添加的抗原在其中的作用 相互作用尚未确定。第一个特定目标使用两个 确定CD1a特异性T细胞是否识别抗原的策略 与CD1a结合或仅识别CD1a的不同部分。第一，氨基 CD1a形成的抗原结合沟底板上的酸 将通过定点突变来改变，以取代任何 肽通常结合在他的凹槽中。如果一个或两个氨基酸的变化 改变多个T细胞克隆对CD1a的识别，然后 假设TCRs与CD1a结合的多肽抗原相互作用将是 支持，因为这些氨基酸不应直接访问 与TCR联系。接下来，来自致敏捐赠者的T细胞将被 筛选识别表达细胞呈递的特定抗原 转染人CD1a，但结合经典MHC分子。只起反应的T细胞 在同时存在CD1a和抗原的情况下，必须识别抗原 由CD1a提交。对于第二个具体目标，估计的数量 可由CD1a呈递的抗原将通过扩增和 CD1a特异性T细胞TCR基因序列测定第三 特定的目的将测试CD1a呈递特定抗原的能力 通过开发表达人CD1a的转基因小鼠来实现活体 小鼠I类MHC基因调控序列。因为转基因小鼠可以 诱导表达具有功能性的人类I类MHC分子 预计转基因人CD1a也将具有功能。老鼠 将接受免疫并测试使用CD1a作为抗原的能力 呈现分子。因为特定抗原的呈递是由非 人类分子的多态之前还没有被证明，这些 研究将提供关于TCR识别抗原和 关于表皮LC的特殊性质。

This project will investigate cutaneous antigen presenting cells by determining the function of CD1a, a non-polymorphic molecule expressed by Langerhans cells (LC). Similarities between CD1a and class I major histocompatibility (MHC) molecules suggest that CD1a can bind peptide antigen. Like class I MHC molecules, mediates cytotoxicity via the T-cell receptor (TCR) for antigen, but the role of added antigen in this interaction has not been determined. The first specific aim uses two strategies to determine whether CD1a specific T-cells recognize antigen bound to CD1a or recognize only distinct portions of CD1a. First, amino acids in the floor of the putative antigen binding groove formed by CD1a will be changed by site directed mutagenesis in order to displace any peptide normally bound int he groove. If changes of one or two amino acids alter the recognition of CD1a by multiple T-cell clones, then the hypothesis that TCRs interact with peptide antigen bound to CD1a will be supported since these amino acids should not be accessible for direct contact with the TCR. Next, T-cells from sensitized donors will be screened for recognition of specific antigens presented by cells expressing transfected CD1a but lacing classical MHC molecules. T-cells reacting only in the presence of both CD1a and antigen must be recognizing the antigen presented by CD1a. For the second specific aim, an estimate of the number of antigens which could be presented by CD1a will be made by amplifying and sequencing TCR mRNA sequences from CD1a specific T-cells. The third specific aim will test the ability of CD1a to present specific antigens in vivo by developing transgenic mice that express human CD1a controlled by murine class I MHC gene regulatory sequences. Since transgenic mice can be induced to express functional human class I MHC molecules, it is anticipated that transgenic human CD1a will also be functional. The mice will be immunized and tested for the ability to use CD1a as an antigen presenting molecule. Because presentation of specific antigens by non- polymorphic human molecules has not been previously demonstrated, these studies will provide new information about TCR recognition of antigens and about special properties of epidermal LC.