Clathrin-mediated Endocytosis of the β-Adrenergic Receptor Is Regulated by Phosphorylation/Dephosphorylation of β-Arrestin1*
- Fang-Tsyr Lin,
- Kathleen M. Krueger,
- Humphrey E. Kendall,
- Yehia Daaka,
- Zoey L. Fredericks,
- Julie A. Pitcher and
- Robert J. Lefkowitz‡
- From the Howard Hughes Medical Institute, Departments of Medicine and Biochemistry, Duke University Medical Center, Durham, North Carolina 27710
Abstract
β-Arrestins serve a dual regulatory role in the life cycle of G protein-coupled receptors such as the β2-adrenergic receptor. First, they mediate rapid desensitization by binding to G protein-coupled receptor kinase-phosphorylated receptors. Second, they target the receptors for internalization into endosomal vesicles, wherein receptor dephosphorylation and resensitization occur. Here we report that phosphorylation of a carboxyl-terminal serine (Ser-412) in β-arrestin1 regulates its endocytotic but not its desensitization function. Cytoplasmic β-arrestin1 is constitutively phosphorylated and is recruited to the plasma membrane by agonist stimulation of the receptors. At the plasma membrane, β-arrestin1 is rapidly dephosphorylated, a process that is required for its clathrin binding and receptor endocytosis but not for its receptor binding and desensitization. Once internalized, β-arrestin1 is rephosphorylated. Thus, as with the classical endocytic adaptor protein complex AP2, β-arrestin1 functions as a clathrin adaptor in receptor endocytosis which is regulated by dephosphorylation at the plasma membrane.
Footnotes
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↵* This work was supported by the Howard Hughes Medical Institute and Grant HL16037 from the National Institutes of Health.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence should be addressed: Howard Hughes Medical Institute, Depts. of Medicine and Biochemistry, Duke University Medical Center, Box 3821, Durham, NC 27710. Tel.: 919-684-2974; Fax: 919-684-8875.
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↵1 The abbreviations used are: kb, kilobase pair; PAGE, polyacrylamide gel electrophoresis; HPLC, high pressure liquid chromatography; β2-AR, β2-adrenergic receptors; PCR, polymerase chain reaction; PBS, phosphate-buffered saline; PAGE, polyacrylamide gel electrophoresis; PVDF, polyvinylidene difluoride.
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↵2 H. Attramadal and R. J. Lefkowitz, unpublished data.
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- Received August 8, 1997.
- Revision received September 24, 1997.
