Effect of 5-bromodeoxyuridine on the induction of adenosine 3′:5′-monophosphate phosphodiesterase in 3T3-L fibroblasts

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Cyclic adenosine 3′:5′-monophosphate (cAMP) phosphodiesterase activity is increased threefold in 3T3-L fibroblasts by 1-methyl-3-isobutylxanthine. Actinomycin D (2 μg/ml) and cycloheximide (10 μg/ml) prevent this increase. Cells pretreated with inducer (0.5 m m) in the presence of cycloheximide for 12 h, rinsed, and then treated with actinomycin D ( t = 0) show a nearly twofold increase in enzyme activity in the next 90 min. The results suggest that cAMP phosphodiesterase is an inducible enzyme in 3T3-L cells and that the increase in enzyme activity requires transcription. The induction of cAMP phosphodiesterase was employed as a model to investigate enzyme induction in cells grown in the presence of the thymidine analog, 5-bromodeoxyuridine. Growth of 3T3-L fibroblasts in 1 μ m bromodeoxyuridine results in an inhibition of the induction of cAMP phosphodiesterase by 1-methyl-3-isobutylxanthine (0.5 m m). This effect increases when bromodeoxyuridine is raised from 0.1 to 10 μ m. The basal level of the enzyme is not altered by bromodeoxyuridine. Addition of thymidine (0.1 to 10 μ m) to the culture medium prevents the bromodeoxyridine effect, indicating that the analog must be incorporated into DNA to act. An increase in the inducer concentration from 0.1 to 1.0 m m partially overcame the bromodeoxyuridine effect on enzyme induction. These results suggest that the induction of cAMP phosphodiesterase can be inhibited by the incorporation of bromodeoxyuridine into DNA, and that an inducer of the enzyme can override the inhibition but at higher concentrations than are necessary in control cultures.