CHARACTERIZATION OF THREE NEW T LYMPHOCYTE-SPECIFIC GENE

我们开发了一个协议，使我们能够隔离广泛的 T淋巴细胞亚群特异性cDNA的代表，和 将该方法应用于ConA刺激的克隆Th 和CTL cDNA文库。 16种以前未识别的T细胞 已经分离出了特定的基因，除了七个已知的T- 细胞基因 16个基因中有3个在两种Th细胞中都表达， 和CTL，7个仅在Th中表达，6个仅在CTL中表达。 16个基因进一步表征， 用ConA-、IL-2、T细胞处理后的表达模式 受体抗体或环孢菌素A。 Nuclotide序列分析 发现了其中五个基因， 描述了已知和未知功能的基因。 客观 本建议的目的是展示三个功能， 选择上述基因是因为它们是可诱导的， ConA优先在Th中表达，似乎代表了新的可溶性 T细胞介质。 该策略将是制备抗体， 每种基因产物识别相应的 使用寡肽或E.大肠杆菌重组蛋白 作为抗原。 接下来，基因产物将在 从真核表达系统中纯化，其可以模拟 接近天然产物。 针对每种抗体的抗体 三种cDNA产物和纯化的重组蛋白 然后将被用来确定每一个的生物功能， 基因产物 计划的研究包括受体结合 研究来确定哪些细胞携带这些分子的受体， 并使用各种免疫学方法证明其功能， 测定。 为了补充体外免疫学测定， 将通过以下方法筛选具有各种免疫学病症的突变体： cDNA探针来鉴定这些突变体是否携带异常 对应的基因。 长期目标是确定 每个分子的人类同源物， 人类免疫缺陷和其他疾病的临床应用 例如人类恶性肿瘤和艾滋病。 本 调查可以为示范提供一个模型研究 功能和临床应用的未知分子， 已经在核酸水平上得到了确认。

We have developed a protocol which allowed us to isolated broad representation of T-lymphocyte subset-specific cDNAs, and applied the method to the analysis of ConA-stimulated cloned Th and CTL cDNA libraries. Sixteen previously unrecognized T cell specific genes have been isolated, in addition to seven known T- cell genes. Three of the sixteen genes were expressed in both Th and CTL, seven were expressed in only Th and six only in CTL. The 16 genes were further characterized according to the expression pattern after treatment with ConA-, IL-2, T-cell receptor antibody or cyclosporin A. Nuclotide sequence analyses identified five of these genes as corresponding to recently described genes of known and unknown functions. The objective of the present proposal is to demonstrate the functions of three of the above genes which were selected because they are inducible by ConA preferentially in Th and appear to represent new soluble T-cell mediators. The strategy will be to prepare antibodies to each of the gene products which recognize the corresponding native proteins using oligopeptides or E. coli recombinant proteins as antigens. Next, the gene products will be produced in the purified from eukaryotic expression systems, which may simulate closely the natural product. The antibodies specific to each of the three cDNA products and the purified recombinant proteins will then be utilized to determine the biologic function(s) of each gene product. The investigations planned include receptor binding studies to identify which cells carry receptors to these molecules, and to demonstrate their functions using various immunological assays. To supplement the in vitro immunological assays, mouse mutants with various immunological disorders will be screened by the cDNA probes to identify if such mutants carry abnormalities of the corresponding genes. A long term objective is to identify the human homologue of each of the molecules and to seek clinical applications for human immunodeficiency and other diseases such as human malignancies and AIDS. The present investigation could present a model study for the demonstration of functions and a clinical applications of unknown molecules that have been identified at the nucleic acid level.