Weiser W Y, Temple P A, Witek-Giannotti J S, Remold H G, Clark S C, David J R
Department of Medicine, Harvard Medical School, Boston, MA 02115.
Proc Natl Acad Sci U S A. 1989 Oct;86(19):7522-6. doi: 10.1073/pnas.86.19.7522.
A cDNA encoding a human macrophage migration inhibitory factor (MIF) was isolated, through functional expression cloning in COS-1 cells, from a cDNA library prepared from a lectin-stimulated T-cell hybridoma, T-CEMB. The 115-amino acid polypeptide encoded by the MIF cDNA (p7-1) was effectively released from the transfected COS-1 cells and yielded readily detectable MIF activity in the culture supernatant despite the apparent lack of a classical protein secretory sequence. Insertional mutational analysis and elution of MIF activity from polyacrylamide gel slices demonstrated that the Mr 12,000 protein with MIF activity released by the COS-1 cells is encoded by p7-1. The p7-1 cDNA hybridized with a 700-base mRNA expressed by Con-A-stimulated lymphocytes but not unstimulated lymphocytes. The availability of the MIF cDNA clone and recombinant MIF will facilitate the analysis of the role of this lymphokine in cell-mediated immunity, immunoregulation, and inflammation.
通过在COS-1细胞中进行功能表达克隆，从由凝集素刺激的T细胞杂交瘤T-CEMB制备的cDNA文库中分离出了编码人巨噬细胞移动抑制因子（MIF）的cDNA。MIF cDNA（p7-1）编码的115个氨基酸的多肽能有效地从转染的COS-1细胞中释放出来，尽管明显缺乏经典的蛋白质分泌序列，但在培养上清液中仍能产生易于检测到的MIF活性。插入突变分析以及从聚丙烯酰胺凝胶切片上洗脱MIF活性表明，COS-1细胞释放的具有MIF活性的12,000 Mr蛋白由p7-1编码。p7-1 cDNA与伴刀豆球蛋白A刺激的淋巴细胞而非未刺激的淋巴细胞所表达的700个碱基的mRNA杂交。MIF cDNA克隆和重组MIF的可得性将有助于分析这种淋巴因子在细胞介导的免疫、免疫调节和炎症中的作用。
