Stable interaction between beta-arrestin 2 and angiotensin type 1A receptor is required for beta-arrestin 2-mediated activation of extracellular signal-regulated kinases 1 and 2.
Binding of beta-arrestins to seven-membrane-spanning receptors (7MSRs) not only leads to receptor desensitization and endocytosis but also elicits additional signaling processes. We recently proposed that stimulation of the angiotensin type 1A (AT(1A)) receptor results in independent beta-arrestin 2- and G protein-mediated extracellular signal-regulated kinases 1 and 2 (ERK1/2) activation. Here we utilize two AT(1A) mutant receptors to study these independent pathways, one truncated at residue 324, thus removing all potential carboxyl-terminal phosphorylation sites, and the other bearing four mutations in the serine/threonine-rich clusters in the carboxyl terminus. As assessed by confocal microscopy, the two mutant receptors interacted with beta-arrestin 2-green fluorescent protein with much lower affinity than did the wild-type receptor. In addition, the mutant receptors more robustly stimulated G protein-mediated inositol phosphate production. Approximately one-half of the wild-type AT(1A) receptor-stimulated ERK1/2 activation was via a beta-arrestin 2-dependent pathway (suppressed by beta-arrestin 2 small interfering RNA), whereas the rest was mediated by a G protein-dependent pathway (suppressed by protein kinase C inhibitor). ERK1/2 activation by the mutant receptors was insensitive to beta-arrestin 2 small interfering RNA but was reduced more than 80% by a protein kinase C inhibitor. The biochemical consequences of ERK activation by the G protein and beta-arrestin 2-dependent pathways were also distinct. G-protein-mediated ERK activation enhanced the transcription of early growth response 1, whereas beta-arrestin 2-dependent ERK activation did not. In addition, stimulation of the truncated AT(1A) mutant receptor caused significantly greater early growth response 1 transcription than did the wild-type receptor. These findings demonstrate how the ability of receptors to interact with beta-arrestins determines both the mechanism of ERK activation as well as the physiological consequences of this activation.
Wei
Huijun
H
Howard Hughes Medical Institute, Departments of Medicine and Biochemistry, Duke University Medical Center, Durham, North Carolina 27710, USA.
Ahn
Seungkirl
S
Barnes
William G
WG
Lefkowitz
Robert J
RJ
D000595:Amino Acid Sequence; D000804:Angiotensin II / Q000378:metabolism; D000818:Animals; D019390:Arrestins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D051766:Early Growth Response Protein 1; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics / Q000378:metabolism; D007211:Indoles / Q000378:metabolism; D008301:Maleimides / Q000378:metabolism; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D044140:Receptor, Angiotensin, Type 1 / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015290:Second Messenger Systems / Q000502:physiology; D016415:Sequence Alignment; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins
C000605254:ARRB2 protein, human; D019390:Arrestins; D004268:DNA-Binding Proteins; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D017874:Immediate-Early Proteins; D007211:Indoles; D008301:Maleimides; D034741:RNA, Small Interfering; D044140:Receptor, Angiotensin, Type 1; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins; D000804:Angiotensin II; C071924:Ro 31-8425; D011493:Protein Kinase C; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D019204:GTP-Binding Proteins