Regions essential for the expression of the EGF receptor gene have been identified by deleting various portions of the promoter. Proteins which can bind to these regions have been identified by their ability to protect DNA from the promoter region from being digested by exonuclease III (exonuclease mapping). An oligonucleotide corresponding to one of the sites has been synthesized and used to purify one of these proteins; this protein is currently being characterized. A crude cell-free system which carries out transcription of the EGF receptor has been developed. Deletion analysis of the EGF receptor template has identified a small region necessary to support transcription. A crude transcription extract has been subjected to fractionation by heparin agarose and DEAE cellulose chromatography to identify factors necessary for transcription. Methylation of the DNA template appears to have no effect on RNA transcription. Previously, EGF and phorbol ester (PMA) were shown to promote accumulation of EGF receptor mRNA. Currently possible mechanisms responsible for receptor mRNA accumulation are being evaluated to distinguish between effects of these ligands on initiation of transcription and mRNA stabilization.
