Genetic Characterization of a Brain-Specific Form of the Type I Regulatory Subunit of cAMP-Dependent Protein Kinase

An isoform (RIβ ) of the regulatory type I subunit gene of cAMP-dependent protein kinase (EC 2.7.1.37) has been characterized in mouse. The open reading frame of the RIβ cDNA is 72% identical in nucleotide sequence with the previously cloned RI gene, now referred to as RIα . Both genes code for a protein of 380 amino acids and their proteins are 82% identical in amino acid sequence. Sequence similarity is highest in the regions that form the pseudosubstrate-binding site of the catalytic subunit and the two cAMP binding domains. The amino-terminal portion shows the greatest dissimilarity, suggesting that the isoforms may differ in their dimerization properties or interaction with other proteins. In contrast to RIα , which is constitutively expressed in all tissues, RIβ is expressed in a highly tissue-specific manner. Brain and spinal cord contained significant levels of RIβ mRNA, testis RNA gave a detectable signal, and all other tissues tested were negative. Expression of a RIβ cDNA in NIH 3T3 cells resulted in the appearance of a RI subunit protein that migrated more slowly than RIα after NaDodSO4/PAGE. The native form of RIβ in brain could also be distinguished from RIα by its abnormal migration on NaDodSO4/PAGE. RIβ protein produced in 3T3 cells was shown to be functional by its ability to form a cAMP-dependent holoenzyme with the catalytic subunit.