Kitamura T, Sato N, Arai K, Miyajima A
Department of Molecular Biology, DNAX Research Institute of Molecular and Cellular Biology, Palo Alto, California 94304.
Cell. 1991 Sep 20;66(6):1165-74. doi: 10.1016/0092-8674(91)90039-2.
A cDNA for a human interleukin-3 (hIL-3) binding protein has been isolated by a novel expression cloning strategy: a cDNA library was coexpressed with the cDNA for the beta subunit of human granulocyte/macrophage colony-stimulating factor (GM-CSF) receptor (hGMR beta) in COS7 cells and screened by binding of 125I-labeled IL-3. The cloned cDNA (DUK-1) encodes a mature protein of 70 kd, which belongs to the cytokine receptor family and which alone binds hIL-3 with extremely low affinity (Kd = 120 +/- 60 nM). A high affinity IL-3-binding site (Kd = 140 +/- 30 pM) was reconstituted by coexpressing the DUK-1 protein and hGMR beta, indicating that hIL-3R and hGMR share the beta subunit. Therefore, we designated DUK-1 as the alpha subunit of the hIL-3R. As in human hematopoietic cells, hIL-3 and hGM-CSF complete for binding in fibroblasts expressing the cDNAs for hIL-3R alpha, GMR alpha, and the common beta subunit, indicating that different alpha subunits compete for a common beta subunit.
通过一种新型表达克隆策略分离出了一种人白细胞介素-3（hIL-3）结合蛋白的互补DNA（cDNA）：将一个cDNA文库与人粒细胞/巨噬细胞集落刺激因子（GM-CSF）受体（hGMRβ）β亚基的cDNA在COS7细胞中共表达，并用125I标记的IL-3结合进行筛选。克隆的cDNA（DUK-1）编码一个70kd的成熟蛋白，它属于细胞因子受体家族，单独与hIL-3结合时亲和力极低（Kd = 120±60 nM）。通过共表达DUK-1蛋白和hGMRβ重建了一个高亲和力的IL-3结合位点（Kd = 140±30 pM），这表明hIL-3R和hGMR共享β亚基。因此，我们将DUK-1命名为hIL-3R的α亚基。与人类造血细胞一样，hIL-3和hGM-CSF在表达hIL-3Rα、GMRα和共同β亚基cDNA的成纤维细胞中竞争结合，这表明不同的α亚基竞争一个共同的β亚基。
