The exposure of quiescent Swiss 3T3 cells to mainstream cigarette smoke (CS) trapped in PBS solution (smoke-bubbled PBS) resulted in the dose-dependent expression of c-fos mRNA and protein. Kinetic investigations revealed that in contrast to mitogens, which strongly but transiently induce the c-fos promoter within minutes, c-fos transcripts in cells exposed to 0.03 puffs (∼1 cm3) of CS/ml of medium accumulated slowly but were still seen after 8 h; the maximum expression rates were between 2 and 6 h of exposure. This specific expression pattern appears to be the result of altered posttranscriptional as well as transcriptional regulation, since a strikingly increased stability of the c-fos message (t½, ≥2 h versus <20 min in serum-stimulated cells) in smoke-treated cells was observed in addition to slight transcriptional activation of the c-fos promoter. CS-dependent DNA damage can be excluded as the only source for this altered expression pattern, since inhibition of DNA strand break formation by either cataiase or o-phenanthroline had no detectable effect on the CS-induced c-fos expression. The results described here, and other CS-dependent cellular and biochemical effects, are similar to those induced in vitro by okadaic acid, a specific inhibitor of cell growth-regulatory protein phosphatases 1/2A (PP-1/2A). Hence, the effects of smoke treatment on these key enzymes were compared to those of okadaic acid based on the ability of cell-free extracts to release radiolabeled phosphate from glycogen phosphorylase a, a substrate of PP-1/2A. Results from these experiments indicate that both treatments inhibited PP-1/2A in a concentration- and analogous time-dependent manner. The data presented suggest that PP-1/2A may, at least in vitro, be targeted by water-soluble active compounds present in cigarette smoke.