STRUCTURE OF 2 RAT GENES-CODING FOR CLOSELY-RELATED ROLIPRAM-SENSITIVE CAMP PHOSPHODIESTERASES - MULTIPLE MESSENGER-RNA VARIANTS ORIGINATE FROM ALTERNATIVE SPLICING AND MULTIPLE START SITES

The products of two phosphodiesterase (PDE) genes (ratPDE3/IVd and rat PDE4/IVb) are present in the rat Sertoli cell in culture, and their ex pression is under the control of the gonadotropin follicle-stimulating hormone (Swinnen, J. V., Tsikalas, K. E., and Conti, M. (1991) J. Bio l. Chem. 266,18370-18377). To understand the basis of the sequence het erogeneity found in the 5'-region of the different cDNAs thus far char acterized, the structure of the coding region of these two cAMP PDE ge nes was investigated. Analysis of five ratPDE3/IVd and ratPDE4/IVb gen omic clones showed that the coding region of these genes expressed in the Sertoli cell is divided into 11 exons distributed over 35-45 kilob ases of genomic DNA. The intron/exon boundaries agreed, with some exce ptions, with the established consensus sequences and were located in t he same position in the coding region of the two genes. Also present w ere similarities to the exon composition of the Drosophila melanogaste r ''dunce'' gene, the ancestor of these mammalian cAMP PDEs. Multiple AUG codons and short open reading frames were present at the 5'-untran slated end of the ratPDE4/IVb mRNA, but not in the ratPDE3 mRNA. By us ing polymerase chain reaction amplification or Northern analysis, it w as 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 m RNA variants are generated by inclusion or removal of an intron sequen ce that produces a frameshift affecting the position of the initiation AUG codon. Both mRNA species were efficiently translated into cAMP PD E proteins with different molecular masses in a transient transfection assay in COS cells. Polymerase chain reaction amplification demonstra ted that heterogeneity of ratPDE4/IVb mRNAs was present in the same lo cation as in the ratPDE3/IVd mRNA. Two ratPDE4/IVb mRNAs with differen t 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 th at controls the transcription of this mRNA in Sertoli and FRTL-5 cells , but not in the brain. Upstream exons and additional promoters are pr obably present and necessary to generate the brain-specific mRNAs. The se 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.