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The palmitoylation state of the β 2 ‐adrenergic receptor regulates the synergistic action of cyclic AMP‐dependent protein kinase and β‐adrenergic receptor kinase involved in its phosphorylation and desensitization
Author(s) -
Moffett Serge,
Rousseau Guy,
Lagacé Monique,
Bouvier Michel
Publication year - 2001
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2001.00005.x
Subject(s) - homologous desensitization , beta adrenergic receptor kinase , phosphorylation , desensitization (medicine) , agonist , palmitoylation , protein kinase a , receptor , biology , chemistry , kinase , endocrinology , biochemistry , g protein coupled receptor , enzyme , cysteine
Although palmitoylation of the β 2 ‐adrenergic receptor (β 2 AR), as well as its phosphorylation by the cyclic AMP‐dependant protein kinase (PKA) and the β‐adrenergic receptor kinase (βARK), are known to play important roles in agonist‐promoted desensitization, their relative contribution and mutual regulatory influences are still poorly understood. In this study, we investigated the role that the carboxyl tail PKA site (Ser 345,346 ) of the β 2 AR plays in its rapid agonist‐promoted phosphorylation and desensitization. Mutation of this site (Ala 345,346 β 2 AR) significantly reduced the rate and extent of the rapid desensitization promoted by sustained treatment with the agonist isoproterenol. The direct contribution of Ser 345,346 in desensitization was then studied by mutating all other putative PKA and βARK phosphorylation sites (Ala 261,262 βARK − β 2 AR). We found this mutant receptor to be phosphorylated upon receptor activation but not following direct activation of PKA, suggesting a role in receptor‐specific (homologous) but not heterologous phosphorylation. However, despite its phosphorylated state, Ala 261,262 βARK − β 2 AR did not undergo rapid desensitization upon agonist treatment, indicating that phosphorylation of Ser 345,346 alone is not sufficient to promote desensitization. Taken with the observation that mutation of either Ser 345,346 or of the βARK phosphorylation sites prevented both the hyper‐phosphorylation and constitutive desensitization of a palmitoylation‐less mutant (Gly 341 β 2 AR), our data suggest a concerted/synergistic action of the two kinases that depends on the palmitoylation state of the receptor. Consistent with this notion, in vitro phosphorylation of Gly 341 β 2 AR by the catalytic subunit of PKA facilitated further phosphorylation of the receptor by purified βARK. Our study therefore allows us to propose a coordinated mechanism by which sequential depalmitoylation, and phosphorylation by PKA and βARK lead to the functional uncoupling and desensitization of the β 2 AR.