Structure of two rat genes coding for closely related rolipram-sensitive cAMP phosphodiesterases. Multiple mRNA variants originate from alternative splicing and multiple start sites.

The products of two phosphodiesterase (PDE) genes (ratPDE3/IVd and ratPDE4/IVb) are present in the rat Sertoli cell in culture, and their expression is under the control of the gonadotropin follicle-stimulating hormone (Swinnen, J.V., Tsikalas, K.E., and Conti, M. (1991) J. Biol. Chem. 266, 18370-18377). To understand the basis of the sequence heterogeneity found in the 5'-region of the different cDNAs thus far characterized, the structure of the coding region of these two cAMP PDE genes was investigated. Analysis of five ratPDE3/IVd and ratPDE4/IVb genomic clones showed that the coding region of these genes expressed in the Sertoli cell is divided into 11 exons distributed over 35-45 kilobases of genomic DNA. The intron/exon boundaries agreed, with some exceptions, with the established consensus sequences and were located in the same position in the coding region of the two genes. Also present were similarities to the exon composition of the Drosophila melanogaster "dunce" gene, the ancestor of these mammalian cAMP PDEs. Multiple AUG codons and short open reading frames were present at the 5'-untranslated end of the ratPDE4/IVb mRNA, but not in the ratPDE3 mRNA. By using polymerase chain reaction amplification or Northern analysis, it was determined that at least two forms of ratPDE3/IVd mRNA are present in rat Sertoli and FRTL-5 thyroid cells, but not in the brain. These mRNA variants are generated by inclusion or removal of an intron sequence that produces a frameshift affecting the position of the initiation AUG codon. Both mRNA species were efficiently translated into cAMP PDE proteins with different molecular masses in a transient transfection assay in COS cells. Polymerase chain reaction amplification demonstrated that heterogeneity of ratPDE4/IVb mRNAs was present in the same location as in the ratPDE3/IVd mRNA. Two ratPDE4/IVb mRNAs with different 5'-ends were expressed in Sertoli and FRTL-5 cells and in the brain. This heterogeneity is caused by the presence of an intron promoter that controls the transcription of this mRNA in Sertoli and FRTL-5 cells, but not in the brain. Upstream exons and additional promoters are probably present and necessary to generate the brain-specific mRNAs. These findings demonstrate that the cAMP-specific PDE genes have complex structure and that cAMP PDE proteins with different amino termini are derived from these genes.