Several mRNAs coding for a cAMP-specific phosphodiesterase (ratPDE3/IVd) with divergent 5' regions have been detected in mammalian cells. To determine the physiological significance of these differences, the expression of these mRNAs and the properties of the corresponding proteins were investigated. At least three mRNA species derived from the ratPDE3/IVd gene (ratPDE3.1, ratPDE3.2, and ratPDE3.3 mRNAs) are present in Sertoli and thyroid cells and in brain. Expression of ratPDE3.1 and ratPDE3.2 but not ratPDE3.3 mRNAs was dependent on hormone stimulation. The ratPDE3.2 and ratPDE3.3 mRNA variants were translated into polypeptides with immunochemical and biochemical properties identical to the native cAMP phosphodiesterases (PDEs) found in the Sertoli cell and thyroid FRTL-5 cells. Incubation of the recombinant PDEs with the catalytic subunit of the cAMP-dependent protein kinase in a cell-free system caused the phosphorylation and activation of the ratPDE3.3 protein variant. Under the same experimental conditions, ratPDE3.1 and ratPDE3.2 protein variants were neither phosphorylated nor activated by the cAMP-dependent protein kinase. Similar results were obtained by stimulating cells expressing the three recombinant PDE variants with dibutyryl cAMP. These findings demonstrate that the ratPDE3/IVd gene codes for PDE forms subject to different regulations.