Streptogramin-based gene regulation systems for mammalian cells
Streptogramin-based gene regulation systems for mammalian cells
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DOI：
10.1038/81208
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发表时间：
2000-11-01
影响因子：
46.9
通讯作者：
Bailey, JE
中科院分区：
文献类型：
--
作者：
Fussenegger, M;Morris, RP;Bailey, JE
在这里，我们描述了在哺乳动物细胞中调节基因表达的可抑制系统(PIPOFF)和可诱导系统(PIPON)，该系统基于由天蓝色链霉菌对链霉素耐药操纵子编码的阻遏蛋白(PIP)。在这些系统的控制下，基因的表达对临床批准的属于链霉素组的抗生素(原始霉素、维吉尼亚霉素和协菌素)有反应。通过链球菌素调节小鼠促红细胞生成素(EPO)、人胎盘分泌型碱性磷酸酶(SEAP)或绿色荧光蛋白(GFP)在不同细胞系(BHK、CHO、HeLa和小鼠成肌细胞)中的表达，证明了这些系统的多功能性。对CHO细胞中同基因结构的分析表明，与广泛使用的四环素抗性(TetOFF)表达系统相比，PipOFF系统具有更低的本底和更高的诱导率。基于链球菌素的表达技术在功能上与TetOFF系统兼容，从而能够选择性地使用不同的抗生素来独立控制同一细胞中两种不同的基因活动。
Here we describe repressible (PipOFF) as well as inducible (PipON) systems for regulated gene expression in mammalian cells, based on the repressor Pip (pristinamycin-induced protein), which is encoded by the streptogramin resistance operon of Streptomyces coelicolor. Expression of genes placed under control of these systems was responsive to clinically approved antibiotics belonging to the streptogramin group (pristinamycin, virginiamycin, and Synercid). The versatility of these systems was demonstrated by streptogramin-regulated expression of mouse erythropoietin (EPO), human placental secreted alkaline phosphatase (SEAP), or green fluorescent protein (GFP) in diverse cell lines (BHK, CHO, HeLa, and mouse myoblasts). Analysis of isogenic constructs in CHO cells demonstrated the PipOFF system gave lower background and higher induction ratios than the widely used tetracycline-repressible (TetOFF) expression systems. The streptogramin-based expression technology was functionally compatible with the TetOFF system, thus enabling the selective use of different antibiotics to independently control two different gene activities in the same cell.