Molecular analyses of the association of CD4 with two members of the transmembrane 4 superfamily, CD81 and CD82.
Molecular analyses of the association of CD4 with two members of the transmembrane 4 superfamily, CD81 and CD82.
复制标题
CD4 与跨膜 4 超家族的两个成员 CD81 和 CD82 关联的分子分析。
作者：
Toshio Imai;M. Kakizaki;M. Nishimura;O. Yoshie
以前，我们已经表明，CD 81和CD 82，跨膜4超家族的两个成员，形成多分子膜复合物，通过相互关联，并与T细胞中的CD 4或CD 8。在本研究中，我们进一步分析了分子基础的CD 4与CD 81和CD 82的共沉淀实验。首先，我们通过使用由CD 4和CD 2产生的嵌合蛋白来检查与CD 81和CD 82相关的CD 4区域。证实了在转染的细胞中，CD 4而不是CD 2能够与CD 81和CD 82结合。我们发现，CD 4的胞浆区足以使嵌合蛋白共沉淀CD 81，而CD 4的胞浆区和胞外区两者都是它们有效共沉淀CD 82所必需的。我们接着发现，通过使用缺少C-末端31个氨基酸的截短的CD 4或在394和397处的半胱氨酸残基被丝氨酸取代的突变的CD 4，p56 lck结合位点或用棕榈酸的共价修饰对于CD 4与CD 81和CD 82缔合不是必需的。最后，我们发现p56 lck与CD 4的结合强烈抑制了其与CD 81和CD 82的结合。因此，这表明，CD 4至少存在于两种物理状态，一种与p56 lck相关，另一种与CD 81和CD 82相关的p56 lck的不存在或解偶联。
Previously, we have shown that CD81 and CD82, two members of the transmembrane 4 superfamily, form multimolecular membrane complexes by associating with each other and with CD4 or CD8 in T cells. In the present study, we further analyzed the molecular basis of the CD4 association with CD81 and CD82 by co-precipitation experiments. First, we examined the regions of CD4 involved in the association with CD81 and CD82 by employing chimeric proteins generated from CD4 and CD2. It was confirmed that CD4, but not CD2, was capable of binding with CD81 and CD82 in transfected cells. We found that the cytoplasmic region of CD4 was sufficient for the chimeric proteins to co-precipitate CD81, while both the cytoplasmic and extracellular regions of CD4 were required for them to efficiently co-precipitate CD82. We next found, by using truncated CD4 lacking the C-terminal 31 amino acids or mutated CD4 with the cysteine residues at 394 and 397 replaced by serine, that the p56lck binding site or the covalent modification with palmitic acid was not necessary for CD4 to associate with CD81 and CD82. Finally, we found that the binding of p56lck to CD4 strongly inhibited its association with CD81 and CD82. It is, therefore, suggested that CD4 exists at least in two physical states, one associated with p56lck and another associated with CD81 and CD82 in the absence or uncoupling of p56lck.