Robinson W H, Prohaska S S, Santoro J C, Robinson H L, Parnes J R
Department of Medicine, Stanford University School of Medicine, CA 94305, USA.
J Immunol. 1995 Nov 15;155(10):4739-48.
CD6 is a 105/130 kDa monomeric T cell surface glycoprotein that has been shown to play a role in human T cell activation. Recently a partial mouse CD6 cDNA sequence was described. We have isolated full-length cDNA clones including the initiation codon and sequence encoding the full signal peptide, as well as an additional 39 amino acids within the cytoplasmic domain as compared to the previously reported clone. The predicted full-length mouse CD6 protein contains 665 amino acids and has the features of a type I integral membrane protein. The extracellular domain of mouse CD6 is composed of three repeated cysteine-rich domains similar to those in human CD6, mouse and human CD5, and other members of a family of proteins whose prototype is the type I macrophage scavenger receptor. In marked contrast to the previously published human CD6 sequence, the mouse sequence predicts a long cytoplasmic tail that is not closely related to other proteins and possesses two proline-rich motifs containing the SH3-domain binding consensus sequence, three protein kinase C phosphorylation site motifs, nine casein kinase-2 phosphorylation site motifs, and a serine-threonine-rich motif repeated three times. Northern blot analysis revealed that mouse CD6 mRNA is expressed predominantly in thymus, lymph node, and spleen. A polyclonal antiserum was raised against mouse CD6 by gene gun plasmid DNA immunization of rabbits with the mouse CD6 cDNA in an expression vector. In immunofluorescence analysis this polyclonal antiserum positively stained the surface of cells transfected with the mouse CD6 cDNA in an expression vector, as well as most normal mouse thymocytes and peripheral T cells. CD6 protein is expressed on most CD4+CD8+ double-positive and CD4+ or CD8+ single-positive thymocytes, and is expressed at highest levels on mature CD3high thymocytes. The expression of mouse CD6 in thymocytes and peripheral T cells correlates closely with the expression of the related CD5 molecule. The polyclonal rabbit anti-mouse CD6 Abs immunoprecipitated a major polypeptide of 128 kDa from resting and 130 kDa from PMA- and FCS-activated mouse thymocytes and lymph node cells; it is likely that this increase in size upon activation is due to phosphorylation of mouse CD6 as has been described for human CD6. These data demonstrate that mouse thymocytes and T cells express a 130-kDa cell surface protein homologous to human CD6.
CD6是一种105/130 kDa的单体T细胞表面糖蛋白，已被证明在人类T细胞活化中发挥作用。最近报道了小鼠CD6 cDNA的部分序列。我们分离出了全长cDNA克隆，包括起始密码子和编码完整信号肽的序列，以及与先前报道的克隆相比，胞质结构域内另外39个氨基酸。预测的全长小鼠CD6蛋白含有665个氨基酸，具有I型整合膜蛋白的特征。小鼠CD6的胞外结构域由三个重复的富含半胱氨酸的结构域组成，类似于人类CD6、小鼠和人类CD5以及以I型巨噬细胞清道夫受体为原型的蛋白质家族的其他成员。与先前发表的人类CD6序列形成显著对比的是，小鼠序列预测有一个长的胞质尾巴，它与其他蛋白质没有密切关系，并且具有两个富含脯氨酸的基序，包含SH3结构域结合共有序列、三个蛋白激酶C磷酸化位点基序、九个酪蛋白激酶2磷酸化位点基序以及一个富含丝氨酸 - 苏氨酸的基序重复三次。Northern印迹分析显示，小鼠CD6 mRNA主要在胸腺、淋巴结和脾脏中表达。通过用表达载体中的小鼠CD6 cDNA对兔进行基因枪质粒DNA免疫，制备了针对小鼠CD6的多克隆抗血清。在免疫荧光分析中，这种多克隆抗血清对用表达载体中的小鼠CD6 cDNA转染的细胞表面、以及大多数正常小鼠胸腺细胞和外周T细胞进行了阳性染色。CD6蛋白在大多数CD4 + CD8 +双阳性和CD4 +或CD8 +单阳性胸腺细胞上表达，并且在成熟的CD3高胸腺细胞上表达水平最高。小鼠CD6在胸腺细胞和外周T细胞中的表达与相关CD5分子的表达密切相关。多克隆兔抗小鼠CD6抗体从静止的小鼠胸腺细胞和淋巴结细胞中免疫沉淀出一条128 kDa的主要多肽，从经佛波酯（PMA）和胎牛血清（FCS）激活的细胞中沉淀出130 kDa的多肽；激活后这种大小增加可能是由于小鼠CD6的磷酸化，就像人类CD6所描述的那样。这些数据表明，小鼠胸腺细胞和T细胞表达一种与人类CD6同源的130 kDa细胞表面蛋白。
