Recombinant fragment of protein kinase inhibitor blocks cyclic AMP-dependent gene transcription.
Transcriptional regulation by cyclic adenosine monophosphate (cAMP) in mammalian cells could be mediated by a phosphoprotein substrate of the cAMP-dependent protein kinase or, as in prokaryotes, by a cAMP-binding protein. Two synthetic genes that code for an active fragment of the protein inhibitor of this kinase and a mutant inactive fragment were constructed and used to distinguish these alternatives. Transient expression of the active peptide product specifically inhibited the cAMP-stimulated expression of a cotransfected reporter gene by more than 90 percent, whereas the expression of the inactive peptide did not alter cAMP-stimulated gene expression. The results indicate that an active kinase catalytic subunit is a necessary intermediate in the cAMP stimulation of gene transcription.
Grove
J R
JR
Howard Hughes Medical Institute, Harvard Medical School, Boston, MA 02114.
Price
D J
DJ
Goodman
H M
HM
Avruch
J
J
D000123:Acetyltransferases / Q000235:genetics; D002352:Carrier Proteins / Q000494:pharmacology*; D015500:Chloramphenicol O-Acetyltransferase; D003488:Cyanogen Bromide; D000242:Cyclic AMP / Q000494:pharmacology*; D004274:DNA, Recombinant; D004926:Escherichia coli / Q000235:genetics; D047908:Intracellular Signaling Peptides and Proteins; D009693:Nucleic Acid Hybridization; D010446:Peptide Fragments / Q000494:pharmacology*; D010766:Phosphorylation; D010957:Plasmids; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000494:pharmacology*; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection
D002352:Carrier Proteins; D004274:DNA, Recombinant; D047908:Intracellular Signaling Peptides and Proteins; D010446:Peptide Fragments; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D011994:Recombinant Proteins; C015482:protein kinase modulator; D000242:Cyclic AMP; D000123:Acetyltransferases; D015500:Chloramphenicol O-Acetyltransferase; D011494:Protein Kinases; D003488:Cyanogen Bromide