NCI CLINICAL INVESTIGATOR AWARD

绵羊红细胞受体（又称p50、T11或CD 2）是一种T 淋巴细胞特异性细胞表面糖蛋白， 抗原、凝集素和辅助细胞非依赖性T淋巴细胞 activation. 抗CD 2抗体已被证明 以IL-2依赖方式诱导或阻断T细胞活化。 申办方实验室的目标是克隆CD 2， 研究这一重要的结构-功能关系 分子。 为此，我们分离了5个重组cDNA， 从lambda GT-11文库插入，其（a）范围从1000- 1500 bp，（B）通过Southern分析是交叉反应性的，（c）编码 与多价抗CD 2抗血清反应的蛋白决定簇 和（d）与仅在大肠杆菌中发现的1.7和1.3kb RNA转录物杂交， CD 2阳性T细胞，而非CD 2阴性B或T细胞。 这 将应用该奖项来验证我们的cDNA分离物的身份 通过检测COS-7成纤维细胞中的CD 2表位， 用pcEXV-3-CD 2构建体转染。 该载体含有 SV 40早期启动子和增强子序列。 我们比计划 对CD 2阳性和阴性白血病淋巴细胞的协同作用 用pSV-2-neo和pcEXV-3-CD 2构建体构建的细胞系。 我们的目标 （a）分离表达细胞的稳定转染子， 表面CD 2，（B）通过完整CD 2的FACS分析证明 表位的测定和（c）转染细胞中表位的测定 对抗CD 2抗体、凝集素和免疫球蛋白的CD 2依赖性应答 有丝分裂原 如果我们成功地分离出 功能性CD 2，我们建议未来的实验（a）映射 功能上重要的CD 2表位，如T112和T113，（B） 通过选择性诱变确定涉及哪些结构 在CD 2介导的信号转导中，（c）如果特异性 跨膜突变可以改变CD 2的功能， 对于p185（neu蛋白）和（d）转染子是否表现出IL-2 依赖性CD 2介导的反应性。

The sheep erythrocyte receptor (aka p50, T11 or CD2) is a T lymphocyte specific cell surface glycoprotein which mediates antigen, lectin and accessory cell independent T lymphocyte activation. Anti-CD2 antibodies have been shown to either induce, or block T cell activation in an IL-2 dependent manner. The goal of the sponsors laboratory is to clone CD2 in order to study the structure-function relationship of this important molecule. To that end, we have isolated 5 recombinant cDNA inserts from a lambda GT-11 library which (a) range from 1000- 1500 bp, (b) are cross-reactive by Southern analysis, (c) code for protein determinants reactive with a polyvalent anti-CD2 antisera and (d) hybridize to 1.7 and 1.3 kb RNA transcripts found only in CD2 positive T cells but not CD2 negative B or T cells. This award will be applied to verify the identity of our cDNA isolates as CD2 by detecting CD2 epitopes in COS-7 fibroblasts transfected with pcEXV-3-CD2 constructs. This vector contains the SV40 early promotor and enhancer sequences. We than plan on cotransfecting CD2 positive and negative leukemic lymphoid cell lines with pSV-2-neo and pcEXV-3-CD2 constructs. Our aims are (a) the isolation of stable transfectants which express cell surface CD2, (b) demonstration by FACS analysis of intact CD2 epitopes in transfected cells and (c) assay of transfectants for CD2 dependent responsiveness to anti-CD2 antibodies, lectins and mitogens. If we are successful in isolating transfectants with functional CD2, we propose future experiments to (a) map functionally important CD2 epitopes such as T112 and T113, (b) determine by selective mutagenesis which structures are involved in CD2 mediated signal transduction, (c) if specific transmembrane mutations can alter the function of CD2 as seen for p185 (neu protein) and (d) whether transfectants exhibit IL-2 dependent CD2 mediated responsiveness.