Kageyama R, Merlino G T, Pastan I
Laboratory of Molecular Biology, National Cancer Institute, Bethesda, MD 20892.
Proc Natl Acad Sci U S A. 1988 Jul;85(14):5016-20. doi: 10.1073/pnas.85.14.5016.
We have developed an in vitro transcription system for the epidermal growth factor receptor (EGFR) oncogene by using nuclear extracts of A431 human epidermoid carcinoma cells, which overproduce EGFR. We found that a nuclear factor, termed EGFR-specific transcription factor (ETF), specifically stimulated EGFR transcription by 5- to 10-fold. In this report, ETF, purified by using sequence-specific oligonucleotide affinity chromatography, is shown by renaturing material eluted from a NaDodSO4/polyacrylamide gel to be a protein with a molecular mass of 120 kDa. ETF binds to the promoter region, as measured by DNase I "footprinting" and gel-mobility-shift assays, and specifically stimulates the transcription of the EGFR gene in a reconstituted in vitro transcription system. These results suggest that ETF could play a role in the overexpression of the cellular oncogene EGFR.
我们利用过量表达表皮生长因子受体(EGFR)的A431人表皮样癌细胞的核提取物,开发了一种用于EGFR癌基因的体外转录系统。我们发现一种名为EGFR特异性转录因子(ETF)的核因子可将EGFR转录特异性地刺激5至10倍。在本报告中,通过使用序列特异性寡核苷酸亲和色谱法纯化的ETF,经从NaDodSO4/聚丙烯酰胺凝胶洗脱的复性材料显示为一种分子量为120 kDa的蛋白质。通过DNase I“足迹”和凝胶迁移率变动分析测定,ETF与启动子区域结合,并在重组的体外转录系统中特异性刺激EGFR基因的转录。这些结果表明ETF可能在细胞癌基因EGFR的过表达中起作用。