IDENTIFICATION OF POSITIVE AND NEGATIVE REGULATORY ELEMENTS INVOLVED IN THE RETINOIC ACID CAMP INDUCTION OF FGF-3 TRANSCRIPTION IN F9 CELLS

The proto-oncogene Fgt-3 has been implicated as an important signallin g molecule in vertebrate development. In the mouse, it is expressed fo r a limited time at a multitude of sites from embryonic day 7 to birth . Transcription of Fgf-3 initiates at three promoter regions resulting in the generation of various mRNAs which nevertheless all encode the same protein products. A 1.7kb DNA fragment which encompasses these re gions was joined to the CAT reporter gene and shown to function as a p romoter in embryonal carcinoma cells. In stable transfectants the prom oter retains its retinoic acid inducibility, initiating transcription at the same cap-sites as the endogenous gene. In differentiated F9 cel ls, transient transfection of progressive and targeted deletion mutant s of the promoter region has revealed at least two positive and three negative regulatory elements. With one exception, loss of these elemen ts was shown to dramatically affect promoter activity in stable transf ectants of F9 cells. However the promoter remained inducible by retino ic acid to differing degrees, apart from deletions encompassing PS-4A which essentially abolished promoter activity in both undifferentiated and differentiated cells. The sequences of these potential regulatory regions were further defined using DNase-I footprinting, revealing so me similarities to consensus binding sites for known transcription fac tors.