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Biochem Biophys Res Commun. 2017 May 27;487(2):438-443. doi: 10.1016/j.bbrc.2017.04.083. Epub 2017 Apr 17.

β-arrestins negatively control human adrenomedullin type 1-receptor internalization.

Author information

1
Frontier Science Research Center, University of Miyazaki, Miyazaki 889-1692, Japan. Electronic address: kuwasako@med.miyazaki-u.ac.jp.
2
Division of Circulation and Body Fluid Regulation, Faculty of Medicine, University of Miyazaki, Miyazaki 889-1692, Japan.
3
Noto Marine Laboratory, Institute of Nature and Environmental Technology, Division of Marine Environmental Studies, Kanazawa University, Ishikawa 927-0553, Japan.
4
Frontier Science Research Center, University of Miyazaki, Miyazaki 889-1692, Japan.
5
Department of Pharmacology, Hirosaki University, Graduate School of Medicine, Hirosaki 036-8562, Japan.
6
Department of Molecular and Medical Pharmacology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan.

Abstract

Adrenomedullin (AM) is a potent hypotensive peptide that exerts a powerful variety of protective effects against multiorgan damage through the AM type 1 receptor (AM1 receptor), which consists of the calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 2 (RAMP2). Two β-arrestin (β-arr) isoforms, β-arr-1 and β-arr-2, play a central role in the agonist-induced internalization of many receptors for receptor resensitization. Notably, β-arr-biased agonists are now being tested in phase II clinical trials, targeting acute pain and acute heart failure. Here, we examined the effects of β-arr-1 and β-arr-2 on human AM1 receptor internalization. We constructed a V5-tagged chimera in which the cytoplasmic C-terminal tail (C-tail) of CLR was replaced with that of the β2-adrenergic receptor (β2-AR), and it was transiently transfected into HEK-293 cells that stably expressed RAMP2. The cell-surface expression and internalization of the wild-type or chimeric receptor were quantified by flow cytometric analysis. The [125I]AM binding and the AM-induced cAMP production of these receptors were also determined. Surprisingly, the coexpression of β-arr-1 or -2 resulted in significant decreases in AM1 receptor internalization without affecting AM binding and signaling prior to receptor internalization. Dominant-negative (DN) β-arr-1 or -2 also significantly decreased AM-induced AM1 receptor internalization. In contrast, the AM-induced internalization of the chimeric AM1 receptor was markedly augmented by the cotransfection of β-arr-1 or -2 and significantly reduced by the coexpression of DN-β-arr-1 or -2. These results were consistent with those seen for β2-AR. Thus, both β-arrs negatively control AM1 receptor internalization, which depends on the C-tail of CLR.

KEYWORDS:

Adrenomedullin; Calcitonin receptor-like receptor; Receptor activity-modifying protein 2; Receptor trafficking; Signal transduction; β-arrestins

PMID:
28427767
DOI:
10.1016/j.bbrc.2017.04.083
[Indexed for MEDLINE]

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