Nitric oxide modulates beta(2)-adrenergic receptor palmitoylation and signaling.
To determine whether nitric oxide (NO) modulates the beta-adrenergic signaling pathway, we treated cells expressing beta(2)-adrenergic receptors (beta(2)AR) with the NO donors, 3-morpholinosydnonimine (SIN-1) and 1,2,3,4-oxatriazolium, 5-amino-3-(3-chloro-2-methylphenyl)chloride and determined the intracellular production of cAMP after exposure to beta-adrenergic receptor agonists, cholera toxin and forskolin. NO significantly decreased the potency of the beta-adrenergic agonist, isoproterenol, to stimulate cAMP production without affecting the stimulatory action of forskolin and cholera toxin, which directly activate adenylyl cyclase and G(s), respectively. Treatment with the NO donor increased the guanyl nucleotide-sensitive high affinity constant for the agonist, isoproterenol, thus suggesting that it reduced functional coupling between the receptor and G(s). Stimulation of endogenous NO production by lipopolysaccharide in RAW 264.7 macrophages also caused a significant increase in the EC(50) for isoproterenol-stimulated cAMP production. SIN-1 treatment also led to a reduction in both basal and isoproterenol-stimulated incorporation of [(3)H]palmitate into the beta(2)AR. Signaling through the nonpalmitoylated, Gly(341)beta(2)AR mutant was unchanged by SIN-1 treatment. Given the link between beta(2)AR palmitoylation and its responsiveness to agonist, these results suggest that the primary action of NO was depalmitoylation of the beta(2)AR resulting in decreased signaling through the beta(2)AR.
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Département de Biochimie and Le Groupe de Recherche sur le Système Nerveux Autonome, Université de Montréal, Montréal, Quebec H3T 1J4, Canada.
Bouvier
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Jones
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D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007545:Isoproterenol / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008981:Molsidomine / Q000031:analogs & derivatives / Q000494:pharmacology; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000096:biosynthesis / Q000502:physiology*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D010084:Oxidation-Reduction; D019308:Palmitic Acid / Q000378:metabolism*; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
D020030:Nitric Oxide Donors; D018343:Receptors, Adrenergic, beta-2; D019308:Palmitic Acid; D009569:Nitric Oxide; C002385:linsidomine; D008981:Molsidomine; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D007545:Isoproterenol