EXPRESSION OF CDNAS FOR 2 ISOFORMS OF THE CATALYTIC SUBUNIT OF CAMP-DEPENDENT PROTEIN-KINASE

Abstract

We have previously reported the cloning of cDNAs coding for two isoforms (termed C.alpha. and C.beta.) of the catalytic (C) subunit of cAMP-dependent protein kinase (Uhler, M., Chrivia, J., and McKnight G. S. (1986) J. Biol. Chem. 261, 15360-15363). We have constructed cDNA expression vectors for the C.alpha. and C.beta. proteins using the mouse metallothionein promoter so that mRNA transcripts from the expression vectors are inducible with Zn. In stable transformants of NIH 3T3 and AtT-20 cells, the induction of both C.alpha. and C.beta. mRNA with Zn increases the cAMP-dependent as well as the cAMP-independent kinase activity. The C.alpha. and C.beta. proteins synthesized from these expression vectors can be detected by Western blot analysis and have slightly different molecular weights as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Interestingly, the levels of RI protein are also shown to increase in cells expressing high levels of the C.alpha. or C.beta. subunits although there is no change in RI mRNA. In contrast, the levels of RII protein are not significantly affected by increasing either C.alpha. or C.beta. expression in these cell lines. We conclude that cells can compensate for the increased levels of either C.alpha. or C.beta. subunits with a corresponding elevation of RI protein implying that both isoforms of C can interact with RI to form a holoenzyme. The relevance of these experimental results to the coordinate regulation of cAMP-dependent protein kinase subunits is discussed.