The function of inducible promoter systems in F9 embryonal carcinoma cells.
Embryonal carcinoma (EC) cells represent an important model for studying the regulation of cellular differentiation during embryonic development and tumor formation. The differentiation of EC cells is associated with changes in the expression of a number of cellular genes, some of which have been implicated directly in the regulation of differentiation. To facilitate further studies of the possible roles of cellular gene products during the differentiation of EC cells, we have used transient transfection assays to compare the function of three promoter systems that direct the conditional expression of recombinant gene constructs. One system employs the mouse mammary tumor virus (MMTV) promoter, which is induced by glucocorticoid hormones. The other two systems are based on chimeric transactivator proteins consisting of the bacterial lac repressor or tet repressor, respectively, fused with a viral transactivation domain. The chimeric proteins function in mammalian cells as sequence-specific activators of transcription that are regulated by either lactose analogs or tetracycline. Transient transfections of mouse F9 EC cells and their differentiated cells with an MMTV promoter-reporter gene construct and a second plasmid encoding the rat glucocorticoid receptor resulted in a dramatic induction of reporter gene expression by glucocorticoid hormone of approximately 200-fold. The conditional expression system based on the tetracycline-responsive transactivator exhibited a similar range of reporter gene expression in response to tetracycline. In contrast, the system based on the lac repressor exhibited a much more limited range of conditional reporter gene expression in our studies. These findings and others discussed in this report suggest that the tetracycline-responsive promoter system may be useful for the conditional expression of recombinant gene constructs in F9 EC cells. Furthermore, data are presented indicating that the human beta-actin promoter should be suitable for stable expression of conditional transactivators, such as the tetracycline-responsive transactivator, in F9 cells before and after differentiation.
Miller
K
K
Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha 68198-6805, USA.
Rizzino
A
A
D000199:Actins / Q000096:biosynthesis*; D000818:Animals; D018236:Carcinoma, Embryonal; D002454:Cell Differentiation; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D004742:Enhancer Elements, Genetic; D015870:Gene Expression; D006367:HeLa Cells; D006801:Humans; D008324:Mammary Tumor Virus, Mouse; D051379:Mice; D010957:Plasmids; D011401:Promoter Regions, Genetic; D051381:Rats; D011965:Receptors, Glucocorticoid / Q000096:biosynthesis*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection; D014407:Tumor Cells, Cultured
D000199:Actins; D011965:Receptors, Glucocorticoid; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D015500:Chloramphenicol O-Acetyltransferase