Opposing effects of phorbol-12-myristate-13-acetate, an activator of protein kinase C, on the signaling of structurally related human dopamine D1 and D5 receptors.
The 'cross-talk' between different types of neurotransmitters through second messenger pathways represents a major regulatory mechanism in neuronal function. We investigated the effects of activation of protein kinase C (PKC) on cAMP-dependent signaling by structurally related human D1-like dopaminergic receptors. Human embryonic kidney 293 (HEK293) cells expressing D1 or D5 receptors were pretreated with phorbol-12-myristate-13-acetate (PMA), a potent activator of PKC, followed by analysis of dopamine-mediated receptor activation using whole cell cAMP assays. Unpredictably, PKC activation had completely opposite effects on D1 and D5 receptor signaling. PMA dramatically augmented agonist-evoked D1 receptor signaling, whereas constitutive and dopamine-mediated D5 receptor activation were rapidly blunted. RT-PCR and immunoblotting analyses showed that phorbol ester-regulated PKC isozymes (conventional: alpha, betaI, betaII, gamma; novel: delta, epsilon, eta, theta) and protein kinase D (PKCmicro) are expressed in HEK293 cells. PMA appears to mediate these contrasting effects through the activation of Ca2+-independent novel PKC isoforms as revealed by specific inhibitors, bisindolylmaleimide I, Gö6976, and Gö6983. The finding that cross-talk between PKC and cAMP pathways can produce such opposite outcomes following the activation of structurally similar D1-like receptor subtypes is novel and further strengthens the view that D1 and D5 receptors serve distinct functions in the mammalian nervous and endocrine systems.
Jackson
Adele
A
Ottawa Health Research Institute, Ottawa Hospital (Civic Campus), and Department of Medicine/Cellular and Molecular Medicine/Psychiatry, University of Ottawa, Ottawa, Ontario, Canada.
Sedaghat
Keyvan
K
Minerds
Kirsty
K
James
Carolyn
C
Tiberi
Mario
M
D015056:1-Methyl-3-isobutylxanthine / Q000494:pharmacology; D015152:Blotting, Northern / Q000379:methods; D015153:Blotting, Western / Q000379:methods; D002227:Carbazoles / Q000494:pharmacology; D002461:Cell Line, Transformed; D000242:Cyclic AMP / Q000378:metabolism; D004298:Dopamine / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004347:Drug Interactions; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D010703:Phorbol Esters / Q000494:pharmacology*; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D017447:Receptors, Dopamine D1 / Q000378:metabolism*; D017448:Receptors, Dopamine D2 / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors; D014162:Transfection / Q000379:methods
D002227:Carbazoles; D004791:Enzyme Inhibitors; D007211:Indoles; D010703:Phorbol Esters; D010726:Phosphodiesterase Inhibitors; D020033:Protein Isoforms; D012333:RNA, Messenger; D017447:Receptors, Dopamine D1; D017448:Receptors, Dopamine D2; C081021:Go 6976; C032974:12-O-retinoylphorbol-13-acetate; D000242:Cyclic AMP; D011493:Protein Kinase C; D015056:1-Methyl-3-isobutylxanthine; D004298:Dopamine