Regulation of receptor fate by ubiquitination of activated beta 2-adrenergic receptor and beta-arrestin.
Although trafficking and degradation of several membrane proteins are regulated by ubiquitination catalyzed by E3 ubiquitin ligases, there has been little evidence connecting ubiquitination with regulation of mammalian G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptor (GPCR) function. Agonist stimulation of endogenous or transfected beta2-adrenergic receptors (beta2ARs) led to rapid ubiquitination of both the receptors and the receptor regulatory protein, beta-arrestin. Moreover, proteasome inhibitors reduced receptor internalization and degradation, thus implicating a role for the ubiquitination machinery in the trafficking of the beta2AR. Receptor ubiquitination required beta-arrestin, which bound to the E3 ubiquitin ligase Mdm2. Abrogation of beta-arrestin ubiquitination, either by expression in Mdm2-null cells or by dominant-negative forms of Mdm2 lacking E3 ligase activity, inhibited receptor internalization with marginal effects on receptor degradation. However, a beta2AR mutant lacking lysine residues, which was not ubiquitinated, was internalized normally but was degraded ineffectively. These findings delineate an adapter role of beta-arrestin in mediating the ubiquitination of the beta2AR and indicate that ubiquitination of the receptor and of beta-arrestin have distinct and obligatory roles in the trafficking and degradation of this prototypic GPCR.
Shenoy
S K
SK
Howard Hughes Medical Institute and Department of Medicine, Duke University Medical Center, Box 3821, Durham, NC 27710, USA.
McDonald
P H
PH
Kohout
T A
TA
Lefkowitz
R J
RJ
D000818:Animals; D019390:Arrestins / Q000378:metabolism*; D019556:COS Cells; D002384:Catalysis; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D003546:Cysteine Endopeptidases / Q000378:metabolism; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D008025:Ligases / Q000378:metabolism; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism; D009154:Mutation; D009687:Nuclear Proteins; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051736:Proto-Oncogene Proteins c-mdm2; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D000071557:beta-Arrestins
D019390:Arrestins; D009097:Multienzyme Complexes; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D018343:Receptors, Adrenergic, beta-2; D011994:Recombinant Proteins; D025801:Ubiquitin; D000071557:beta-Arrestins; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2; D044767:Ubiquitin-Protein Ligases; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D008025:Ligases; D007545:Isoproterenol