Messenger ribonucleic acids for alpha- and beta-isoforms of cyclic adenosine 3',5'-monophosphate-dependent protein kinase subunits present in the anterior pituitary: regulation of RII beta and C alpha gene expression by the cyclic nucleotide and phorbol ester

Abstract

Expression of the genes encoding the alpha- and beta-isoforms of the catalytic (C) and regulatory type I (RI) and type II (RII) subunits of cAMP-dependent protein kinases (PKA) in the anterior pituitary gland of rats was examined by hybridization of specific 32P-labeled cDNA probes to mRNA. C alpha, C beta, RI alpha, RI beta, RII alpha, and RII beta mRNAs were all present in the anterior pituitary gland and cultured cells, but in different proportions; C alpha was the most abundant, and RII alpha was the least. For C alpha, C beta, RI beta, and RII alpha, Northern blot analysis revealed single mRNAs with sizes of 2.4, 4.5, 2.8, and 6.0 kilobases (kb), respectively. For RI alpha, three distinct mRNAs (1.7, 3.2, and 3.7 kb) were identified, and for RII beta, two were found (1.8 and 3.5 kb). Compared to other tissues and species, including ovine and rat pituitary and testis, differences in sizes and relative abundance of mRNAs and/or mRNA subtypes were observed, demonstrating that great diversity exists, which may reflect tissue and species variation in posttranscriptional processing of mRNA transcripts. When rat anterior pituitary cells were cultured in the presence of 8-bromo-cAMP (8-Br-cAMP), RII beta mRNA levels increased in a dose- and time-dependent manner to a maximum of about 4- to 5-fold the basal values after 5 h in the presence of 2 mM 8-Br-cAMP. Under the same conditions, the C alpha mRNA, already at high levels, further increased, but to a lesser extent (1.5-2 times compared to basal abundancy). Cholera toxin (6 nM) reproduced the effects of 8-Br-cAMP on both C alpha and RII beta mRNAs. These effects were completely abolished in the presence of actinomycin-D, suggesting that cAMP enhances RII beta and C alpha gene transcription. On the other hand, cell exposure to the phorbol ester 12-O-tetradecanoyl 13-phorbol acetate, a potent activator of protein kinase-C (PKC), readily induced a dose-dependent actinomycin-D-inhibited increase in the RII beta mRNA content to levels about 2-fold those in control unstimulated cells, whereas it depressed levels of C alpha mRNA by 25%. In conclusion, we have found that the six genes coding for the alpha- and beta-isoforms of PKA subunits are all expressed in the anterior pituitary, and that the expression of two of those genes (RII beta and C alpha) is regulated by cAMP and phorbol ester in a similar or opposite manner.(ABSTRACT TRUNCATED AT 400 WORDS)