Mutation of tyrosine-350 impairs the coupling of the beta 2-adrenergic receptor to the stimulatory guanine nucleotide binding protein without interfering with receptor down-regulation.
Long-term stimulation of the beta 2-adrenergic receptor (beta 2AR) leads to an internalization and degradation of the receptor. This down-regulation of the beta 2AR number contributes to the desensitization of the adenylyl cyclase activity induced by chronic exposure to agonists. It was proposed that two tyrosine residues (Tyr-350 and Tyr-354) located in the cytoplasmic tail of the beta 2AR play a crucial role in agonist-induced down-regulation. In addition to perturbation of the down-regulation, the substitution of these tyrosines for alanines also led to a functional uncoupling of the receptor from Gs [Valiquette et al. (1990) Proc. Natl. Acad. Sci. U.S.A. 87, 5089-5093]. To further characterize the relative contribution of Tyr-350 and Tyr-354 to the receptor interaction with Gs and agonist-promoted down-regulation, both tyrosines were individually replaced by alanines and mutant receptors expressed in CHW cells. We show here that mutation of Tyr-350 but not that of Tyr-354 significantly decreased the ability of the beta 2AR to be functionally coupled to Gs and thereby to stimulate the adenylyl cyclase. Moreover, in contrast to the double tyrosine mutation, neither of the single-point mutations affected the agonist-induced down-regulation pattern. These data suggest that the presence of either Tyr-350 or Tyr-354 is sufficient to maintain normal agonist-induced down-regulation whereas the integrity of Tyr-350 is required for an appropriate coupling to Gs.(ABSTRACT TRUNCATED AT 250 WORDS)
Département de Biochimie, Université de Montréal, Quebec, Canada.
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Valiquette
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H
Bonin
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Bouvier
D000262:Adenylyl Cyclases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D015536:Down-Regulation; D019204:GTP-Binding Proteins / Q000502:physiology*; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011943:Receptors, Adrenergic, beta / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D013329:Structure-Activity Relationship; D014162:Transfection; D014443:Tyrosine
D011943:Receptors, Adrenergic, beta; D014443:Tyrosine; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D007545:Isoproterenol