A protein kinase C cDNA without the regulatory domain is active after transfection in vivo in the absence of phorbol ester.
We constructed mutant protein kinase C (PKC) cDNAs which expressed PKC activity in vivo in the absence of phorbol ester activation. A hybrid PKC gene, PKAC, was constructed by substituting the coding region for the N-terminal 253 amino acids of PKC alpha with the N-terminal 17 amino acids of the cyclic AMP-dependent protein kinase catalytic subunit (PKA). A truncated PKC gene, delta PKC beta, lacking the coding region for amino acid positions 6 to 159 of PKC beta was also constructed. These mutant kinase genes expressed under the control of the SR alpha promoter activated the c-fos gene enhancer in Jurkat cells and initiated maturation of Xenopus laevis oocytes. Phorbol ester binding activity was absent in both constructs but was preserved in another hybrid gene, PKCA, which was composed of the coding region for 1 to 253 amino acids of PKC alpha at the N-terminal side and the coding region for 18 to 350 amino acids of PKA at the C-terminal side. These results indicate that elimination of the regulatory domain of PKC produces constitutively active PKC that can bypass activation by the phorbol ester. delta PKC beta, in synergy with a calcium ionophore, was capable of activating the interleukin 2 promoter, indicating that cooperation of PKC-dependent and calcium-dependent pathways is necessary for activation of the interleukin 2 gene.
Muramatsu
M
M
Department of Molecular Biology, DNAX Research Institute of Molecular and Cellular Biology, Palo Alto, California 94304.
Kaibuchi
K
K
Arai
K
K
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004789:Enzyme Activation / Q000187:drug effects; D005809:Genes, Regulator; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics; D009154:Mutation; D010703:Phorbol Esters / Q000378:metabolism / Q000494:pharmacology; D011493:Protein Kinase C / Q000096:biosynthesis / Q000235:genetics*; D014162:Transfection
D007376:Interleukin-2; D010703:Phorbol Esters; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D011493:Protein Kinase C