This study examined the effects of suppressing c-fos oncogene expression on multidrug resistance (MDR). A2780S human ovarian carcinoma cells with resistance to actinomycin D were isolated and the resultant A2780AD cells exhibited the MDR phenotype. A hammerhead ribozyme designed to cleave fos RNA cloned into the pMAMneo plasmid was transfected into A2780AD cells. Induction of the ribozyme resulted in decreased expression of c-fos, as well as that of the MDR gene (mdr-1), c-jun, and mutant p53. The transformants displayed altered morphology and restored sensitivity to chemotherapeutic agents comprising the MDR phenotype. An anti-mdr ribozyme separately expressed in A2780AD cells efficiently degraded mdr-1 mRNA. However, reversal of the MDR phenotype by the anti-mdr ribozyme occurred one-fourth as rapidly as that induced by the anti-fos ribozyme. These results reinforce the central role played by c-fos in drug resistance through its participation in signal transduction pathways.