Influence of exogenous ras and p53 on p-glycoprotein function in immortalized rodent fibroblasts

The ability of ras oncogenes and mutant p53 to activate reporter gene expression from human and rodent mdr1 gene promoters was described, although functional significance of this finding was unclear. We analyzed the influence of various forms of recombinant human ras and p53 on the mdr1 gene expression and P-glycoprotein (Pgp) function in rodent immortalized fibroblasts. The ras genes, in addition to activation of exogenous human mdr1 gene promoter, caused an increase in (i) expression of endogenous mdrl mRNA, (ii) Pgp activity as determined by flow cytometry analysis of Rhodamine 123 exclusion, and (iii) resistance of cells to the cytotoxic action of colchicine and some other drugs. To elucidate whether the same signalling pathway is responsible for multidrug resistance induced by various oncogenes and protein kinase C (PKC), we tested the effects of v-mos and the PKC agonist 12-O-tetradecanoylphorbol-13-acetate. Similarly to cells transformed by ras, a Rat1 subline transformed by the v-mos oncogene was characterized by decreased drug sensitivity. On the contrary, Rat1 cells treated with the protein kinase C agonist 12-O-tetradecanoylphorbol-13-acetate showed neither increased mdr1 mRNA expression nor stimulation of Pgp function. Introduction by retrovirus-mediated gene transfer of wild-type p53 into Ratl cells or into murine p53-deficient 10(1) and 10(3) cells did not change the Pgp function significantly, whereas in Rat1 cells transformed by activated N-ras or v-mos, expression of wild-type p53 caused partial reversion of oncogene-induced drug resistance. In agreement with previously published results, we observed that in a transient transfection assay a majority of p53 mutants stimulated reporter gene expression from an exogenous mdr1 promoter in murine cells. However, in Rat1, 10(1) and 10(3) cell sublines, which permanently express various p53s as a result of retroviral transfer, we found neither increase in mdrl mRNA expression and activation of the mdr1 gene promoter, nor increased drug resistance. Moreover, in ras- and mos-transformed Rat1 cells some p53 mutants (His-273, Trp-248), similarly to wild-type p53, caused a slight decrease in resistance to colchicine. On the other hand, Tyr-141 p53 decreased drug resistance of Rat1 and Rat/mos but not Rat/ras cells. Weak effects of physiological concentrations of p53 proteins on mdrl gene expression and Pgp function as well as differential influence of various p53 mutants on multidrug resistance in various cell contexts allow us to suppose that p53 may influence mdr1 gene expression through other transcription factors, rather than by its direct interaction with the mdr1 gene. © 1995.