Regulation of the rb gene by normal and mutated ras, tpa and EGF

Complete inactivation of the human retinoblastoma gene is believed to be an essential step in tumorigenesis of several different cancers. Using the plasmid pRbCAT2 that contains the Rb promoter region was tested for its ability to promote transcription of the bacterial chloramphenicol acetyltransferase (CAT) gene in a transient expression assay. This plasmid was co-transfected in a short term transfections with the plasmids pHO6T1 and pHO6N1 that contains the mutant and normal H-ras gene respectively, into the human cell line HeLa, by the calcium phosphate technique. It was found that the mutant H-ras gene enhances the activity of the Rb gene promoter in contrast to the normal H-ras gene that inhibits it. The expression of the CAT gene in stable clones of HeLa cells carrying the promoter of Rb gene after treatment with TPA and EGF respectively, was also investigated, whereas TPA enhanced, EGF had no effect on the activity of the Rb gene promoter.