Characterization of a Ca2+/calmodulin-dependent protein kinase cascade. Molecular cloning and expression of calcium/calmodulin-dependent protein kinase kinase.
Recent studies have demonstrated that Ca2+/calmodulin-dependent protein kinase IV (CaM-kinase IV) can mediate Ca(2+)-dependent regulation of gene expression through the phosphorylation of transcriptional activating proteins. We have previously identified and purified a 68-kDa rat brain CaM-kinase kinase that phosphorylates and increases total and Ca(2+)-independent activities of CaM-kinase IV (Tokumitsu, H., Brickey, D. A., Gold, J., Hidaka, H., Sikela, J., and Soderling, T. R. (1994) J. Biol. Chem. 269, 28640-28647). Using a partial amino acid sequence of the purified brain kinase, a CaM-kinase kinase cDNA was cloned from a rat brain cDNA library. Northern blot analysis showed that CaM-kinase kinase mRNA (3.4 kilobases) was expressed in rat brain, thymus, and spleen. Sequence analyses revealed that the cDNA encoded a 505-amino acid protein, which contained consensus protein kinase motifs and was 30-40% homologous with members of the CaM-kinase family. Expression of the cDNA in COS-7 cells yielded an apparent 68-kDa CaM-binding protein, which catalyzed in vitro activation in the presence of Mg2+/ATP and Ca2+/ CaM of CaM-kinases I and IV but not of CaM-kinase II. Co-expression of CaM-kinase kinase with CaM-kinase IV gave a 14-fold enhancement of cAMP-response element-binding protein-dependent gene expression compared with CaM-kinase IV alone. These results are consistent with the hypothesis that CaM-kinases I and IV are regulated through a unique signal transduction cascade involving CaM-kinase kinase.
Tokumitsu
H
H
Vollum Institute, Oregon Health Sciences University, Portland 97201, USA.
Enslen
H
H
Soderling
T R
TR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000201:enzymology; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D017346:Protein Serine-Threonine Kinases; D051381:Rats; D017386:Sequence Homology, Amino Acid; D018411:Spodoptera; D013379:Substrate Specificity
D018076:DNA, Complementary; D017346:Protein Serine-Threonine Kinases; D054737:Calcium-Calmodulin-Dependent Protein Kinase Kinase; D017871:Calcium-Calmodulin-Dependent Protein Kinases