Novel Ca2+/calmodulin-dependent protein kinase II γ-subunit variants expressed in vascular smooth muscle, brain, and cardiomyocytes

Abstract

Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) γ- subunits were cloned from a porcine aortic smooth muscle cDNA library resulting in identification of alternatively spliced CaM kinase II γ(B)- and γ(C)-subunits and a novel γ-subunit variant predicted to encode a 60.2-kDa polypeptide, which was designated the γ(G)-subunit. A clone predicted to encode a 62.2-kDa γ-subunit, designated as γ(E), was isolated with a variable domain structure similar to a γ(B)-subunit but with a 114- nucleotide insertion in the conserved 'association' domain of CaM kinase II subunits. A full-length γ(E)-subunit construct expressed in COS cells resulted in multimeric CaM kinase II holoenzymes (470 kDa) with activation and autoregulatory properties similar to expressed holoenzymes composed of γ(B)-, γ(C)-, or γ(G)-subunits. Expression of γ(E) and related γ- subunit mRNAs containing the 114-base insertion was documented in porcine tissues by reverse transcriptase-polymerase chain reaction. CaM kinase II subunits containing the 38-amino acid insert were identified by Western analysis of partially purified CaM kinase II from carotid arterial smooth muscle and brain using a sequence-specific anti-peptide antibody. Immunoprecipitations of tissue homogenates indicated a comparatively high level of expression of subunits containing the insert in brain and provided evidence for their co-assembly with other more abundant subunits into CaM kinase II heteromultimers. Our analyses indicate the following patterns of γ-subunit expression: vascular smooth muscle, γ(B) > γ(C) > γ(E,G); heart, γ(B) > γ(E,C) > γ(G); brain, γ(E) and related subunits >> γ(A,B,C,G).