The effect of c-Ha-ras transfection on phosphatidylcholine biosynthesis of the keratinocyte cell line HaCaT was investigated. It was shown that ras-transfection caused a 3-fold increase of choline incorporation into phosphatidylcholine. By investigating the mechanisms underlying this phenomenon, two targets were obtained. First, the choline uptake was elevated by 2-fold in ras-transfected HaCaT cells as compared with untransfected HaCaT cells, and second, the activity of the rate-limiting enzyme of phosphatidylcholine biosynthesis, CTP:phosphocholine cytidylyltransferase, was increased by 43%. Stimulation of HaCaT cells and ras-transfected HaCaT cells with oleate revealed that the increased activity of cytidylyltransferase might be due to a higher level of enzyme. In these experiments, a 75% increase of the specific activity of fully stimulated, membrane-bound cytidylyltransferase was found in ras-transfected HaCaT cells. Choline kinase which has been previously described as a target of ras-transfection in fibroblasts was unaffected.