MOLECULAR-CLONING AND EXPRESSION OF THE MURINE INTERLEUKIN-5 RECEPTOR
MOLECULAR-CLONING AND EXPRESSION OF THE MURINE INTERLEUKIN-5 RECEPTOR
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DOI：
10.1002/j.1460-2075.1990.tb07886.x
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发表时间：
1990-12-01
期刊：
影响因子：
11.4
通讯作者：
TAKATSU, K
中科院分区：
文献类型：
--
作者：
TAKAKI, S;TOMINAGA, A;TAKATSU, K
小鼠白细胞介素-5（IL-5）在B细胞产生IG和嗜酸性粒细胞的增殖和分化中起重要作用。 在这里，我们通过从鼠IL-5依赖性早期B细胞系制备的文库的表达筛选分离了编码鼠IL-5受体的cDNA克隆。 在COS 7细胞中表达cDNA文库，并使用抗IL-5受体单克隆抗体通过淘选进行筛选。 推导的氨基酸序列分析表明，该受体为415个氨基酸组成的糖蛋白（M（r）45 284），包括N端疏水区（17个氨基酸）、糖基化胞外区（322个氨基酸）、单跨膜区（22个氨基酸）和胞质尾区（54个氨基酸）。 用cDNA转染的COS 7细胞表达60 kd蛋白，其以单一类别的亲和力（K（D）= 2 - 10 nM）结合IL-5。用鼠IL-5受体的cDNA转染的FDC-P1细胞显示具有低亲和力（K（D）= 6 nM）和高亲和力（K（D）= 30 pM）的IL-5结合位点的表达，并获得对IL-5增殖的应答性，尽管亲本FDC-P1细胞未显示任何可检测的IL-5结合。 此外，分离了几个编码可溶形式IL-5受体的cDNA克隆。 北方印迹分析显示，在显示鼠IL-5结合位点的细胞系中检测到两种mRNA（5.0 kb和5.8 kb）。对IL-5受体的氨基酸序列的同源性搜索揭示，IL-5受体包含最近鉴定的细胞因子受体家族的共同基序。
Murine interleukin-5 (IL-5) is know to play an essential role in Ig production of B cells and proliferation and differentiation of eosinophils. Here, we have isolated cDNA clones encoding a murine IL-5 receptor by expression screening of a library prepared from a murine IL-5 dependent early B cell line. A cDNA library was expressed COS7 cells and screened by panning with the use of anti-IL-5 receptor monoclonal antibodies. The deduced amino acid sequence analysis demonstrates that the receptor is glycoprotein of 415 amino acids (M(r)45 284), including an N-terminal hydrophobic region (17 amino acids), a glycosylated extracellular domain (322 amino acids), a single transmembrane segment (22 amino acids) and a cytoplasmic tail (54 amino acids). COS7 cells transfected with the cDNA expressed a 60 kd protein that bound IL-5 with a single class of affinity (K(D) = 2 - 10 nM). FDC-P1 cells transfected with the cDNA for murine IL-5 receptor showed the expression of IL-5 binding sites with both low (K(D) = 6 nM) and high affinity (K(D) = 30 pM) and acquired responsiveness to IL-5 for proliferation, although parental FDC-P1 cells did not show any detectable IL-5 binding. In addition, several cDNA clones encoding soluble forms of the IL-5 receptor were isolated. Northern blot analysis showed that two species of mRNAs (5.0 kb and 5.8 kb) were detected in cell lines that display binding sites for murine IL-5. Homology search for the amino acid sequence of the IL-5 receptor reveals that the IL-5 receptor contains a common motif of a cytokine receptor family that is recently identified.