Altin J G, Pagler E B, Parish C R
Division of Cell Biology, John Curtin School of Medical Research, Australian National University, Canberra City, A. C. T.
Eur J Immunol. 1994 Feb;24(2):450-7. doi: 10.1002/eji.1830240228.
Previous studies have reported an association of the cell surface adhesion molecule CD2 with the T cell receptor and with CD45 on mouse and human T lymphocytes. In this study the association of CD2 with cell surface molecules was investigated using cell surface biotinylation of T lymphocytes, coupled with immunoprecipitation using two CD2-specific monoclonal antibodies (mAb) (RM2-5 and 12-15) and analysis by SDS-PAGE. Although both CD2 mAb immunoprecipitated CD2 from lysates of murine lymphocytes, it was found that mAb 12-15, but not RM2-5, co-precipitated two other molecules of 95 and 180 kDa. Subsequent studies revealed that the 95- and 180-kDa molecules were associated with a subspecies of CD2 (approximately 5%) on thymocytes, the antigen-specific T cell line D10, and splenic T cells but not B cells. Two lines of evidence were obtained consistent with the 95- and 180-kDa molecules being the beta and alpha chains of LFA-1. Firstly, an analysis of 12-15 mAb immunoprecipitates on 4-12% gels under reducing and nonreducing conditions shows that the 95- and 180-kDa molecules have a molecular weight and migration pattern identical to LFA-1. Secondly, depletion of LFA-1 from lysates with LFA-1 mAb abolished the ability of CD2 mAb 12-15 to co-precipitate the 95- and 180-kDa molecules, thereby identifying these as the beta and alpha chains of mouse LFA-1, respectively. These results provide evidence for the first time for an association of LFA-1 and CD2 on mouse T lymphocytes, and suggest that the association occurs with an immunologically distinct subspecies of CD2 molecules.
先前的研究报道了细胞表面黏附分子CD2与小鼠和人类T淋巴细胞上的T细胞受体以及CD45之间存在关联。在本研究中，利用T淋巴细胞的细胞表面生物素化技术，结合使用两种CD2特异性单克隆抗体（mAb）（RM2-5和12-15）进行免疫沉淀，并通过SDS-PAGE分析，对CD2与细胞表面分子的关联进行了研究。尽管两种CD2 mAb均能从小鼠淋巴细胞裂解物中免疫沉淀出CD2，但发现mAb 12-15能共沉淀出另外两个分子量分别为95 kDa和180 kDa的分子，而RM2-5则不能。随后的研究表明，这两种分子量为95 kDa和180 kDa的分子与胸腺细胞、抗原特异性T细胞系D10以及脾脏T细胞（而非B细胞）上约5%的CD2亚类相关。获得了两条证据，支持这两种分子量为95 kDa和180 kDa的分子分别是淋巴细胞功能相关抗原-1（LFA-1）的β链和α链。首先，在还原和非还原条件下，对12-15 mAb免疫沉淀产物在4%-12%凝胶上进行分析，结果显示这两种分子量为95 kDa和180 kDa的分子具有与LFA-1相同的分子量和迁移模式。其次，用LFA-1 mAb从裂解物中去除LFA-1后，CD2 mAb 12-15共沉淀这两种分子量为95 kDa和180 kDa分子的能力消失，从而确定它们分别是小鼠LFA-1的β链和α链。这些结果首次为小鼠T淋巴细胞上LFA-1与CD2之间的关联提供了证据，并表明这种关联发生在免疫上不同的CD2分子亚类上。
