Abstract. The effect of retinoic acid (RA) on the adipose conversion of 3T3 cells has been studied. Differentiation of 3T3-L1 cells was initiated by addition of 0.5 mM methylisobutylxanthine, 0.3 μM dexamethasone and 10 μg/ml insulin (MDI) to confluent monolayers of preadipocytes for 48 h. During this time, the cells underwent DNA replication and cell division prior to the expression of adipose specific genes. RA administration had no apparent effect on the rate or extent of cell growth, cell division, or DNA replication. However, RA treatment concomitant with MDI addition inhibited triacylglycerol accumulation (I0.5= 6 nM) and the accumulation of the differentiation-dependent mRNAs encoding the adipocyte lipid-binding protein (ALBP) and stearoyl-CoA desaturase 1 (SCD1). No inhibition occurred with RA addition either prior to or after MDI treatment. Runoff transcription revealed that the inhibitory effects of RA occurred at the level of transcription and were persistent. Cells treated with RA during the MDI regimen did not appreciably transcribe ALBP or SCDl mRNAs several days following RA withdrawal. The effects of RA were specific for differentiation-dependent transcripts; 10-6M RA did not inhibit expression of the mRNAs encoding β-tubulin or glutamine synthase. Examination of immediate-early transcription factor expression during the MDI regimen revealed that RA mediated an elevated, prolonged expression of c-Jun mRNA accompanied by diminished expression of c-Fos and Jun-B mRNAs. Given the previously demonstrated role of transcription factor AP-1 in ALBP gene expression, our results suggest that the initiation of expression of this and other adipocyte-specific genes during adipose conversion is regulated by the relative composition of transcription factor AP-1.