To elucidate the effect of peroxisome proliferators on the signal-transduction pathway, we have compared the effect of ciprofibrate, an hypolipaemic agent, on the overall phosphoprotein level between rat and human well differentiated hepatic derived cell lines. The phosphorylation status of several phosphoproteins in the rat Fao cell line was increased by the drug while no changes were observed in the human HepG2 cell line. In rat Fao cells, this increase, which is concentration and time dependent, can be as much as eightfold for 20-kDa and 22-kDa proteins. Wy-14,643, a non-fibrate molecule and a more potent peroxisome proliferator than ciprofibrate, increased the phosphorylation status of the same phosphoproteins. Peroxisome proliferators may act by activating kinases inactive in control cells, by amplifying kinases already active in control cells or by inactivating phosphatases. The phosphoamino acid residues affected are essentially serine and threonine. This modification of the signal-transduction pathway by the peroxisome proliferators in rodent cells appears to be an early event or an independent mechanism of the peroxisome proliferation. These results support the accumulating evidence that the perturbation of this pathway may be a major cause of the hepatomegaly and the hepatocarcinogenesis induced by peroxisome proliferators in rodent species. In contrast, the lack of phosphorylation changes in the human HepG2 cell line supports the non-toxic effect of peroxisome proliferators also used as hypolipaemic agents in humans.