Novel (Rp)-cAMPS analogs as tools for inhibition of cAMP-kinase in cell culture. Basal cAMP-kinase activity modulates interleukin-1 beta action.
Novel (Rp)-cAMPS analogs differed widely in ability to antagonize cAMP activation of pure cAMP-dependent protein kinase I and II and to antagonize actions of cAMP on gene expression, shape change, apoptosis, DNA replication, and protein phosphorylation in intact cells. These differences were related to different abilities of the analogs to stabilize the holoenzyme form relative to the dissociated form of cAMP kinase type I and II. (Rp)-8-Br-cAMPS and (Rp)-8-Cl-cAMPS were the most potent cAMP antagonists for isolated type I kinase and for cells expressing mostly type I kinase, like IPC-81 leukemia cells, fibroblasts transfected with type I regulatory subunit (RI), and primary hepatocytes. It is proposed that (Rp)-8-Br-cAMPS or (Rp)-8-Cl-cAMPS should replace (Rp)-cAMPS as the first line cAMP antagonist, particularly for studies in cells expressing predominantly type I kinase. The phosphorylation of endogenous hepatocyte proteins was affected oppositely by (Rp)-8-Br-cAMPS and increased cAMP, indicating that (Rp)-8-Br-cAMPS inhibited basal cAMP-kinase activity. The inhibition of basal kinase activity was accompanied by enhanced DNA replication, an effect which could be reproduced by microinjected mutant cAMP-subresponsive RI. It is concluded that the basal cAMP-kinase activity exerts a tonic inhibition of hepatocyte replication. (Rp)-8-Br-cAMPS and microinjected RI also desensitized hepatocytes toward inhibition of DNA synthesis by interleukin-1 beta. This indicates that basal cAMP-kinase activity can have a permissive role for the action of another (interleukin-1 beta) signaling pathway.
Gjertsen
B T
BT
Department of Anatomy and Cell Biology, University of Bergen, Norway.
Mellgren
G
G
Otten
A
A
Maronde
E
E
Genieser
H G
HG
Jastorff
B
B
Vintermyr
O K
OK
McKnight
G S
GS
Døskeland
S O
SO
D016475:3T3 Cells; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011905:Genes, ras; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D007700:Kinetics; D008099:Liver / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D051379:Mice; D051381:Rats; D017208:Rats, Wistar; D013329:Structure-Activity Relationship; D013873:Thionucleotides / Q000494:pharmacology*; D013936:Thymidine / Q000378:metabolism; D014407:Tumor Cells, Cultured
D007375:Interleukin-1; D013873:Thionucleotides; D005576:Colforsin; C016957:adenosine-3',5'-cyclic phosphorothioate; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D013936:Thymidine