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Activity-Dependent Internalization of Smoothened Mediated by ß-Arrestin 2 and GRK2
Wei Chen,1*Xiu-Rong Ren,2Christopher D. Nelson,2Larry S. Barak,3James K. Chen,4Philip A. Beachy,4Frederic de Sauvage,5Robert J. Lefkowitz2*
Binding of Sonic Hedgehog (Shh) to Patched (Ptc) relieves thelatter's tonic inhibition of Smoothened (Smo), a receptor thatspans the cell membrane seven times. This initiates signalingwhich, by unknown mechanisms, regulates vertebrate developmentalprocesses. We find that two molecules interact with mammalianSmo in an activation-dependent manner: G protein–coupledreceptor kinase 2 (GRK2) leads to phosphorylation of Smo, andß-arrestin 2 fused to green fluorescent protein interactswith Smo. These two processes promote endocytosis of Smo inclathrin-coated pits. Ptc inhibits association of ß-arrestin2 with Smo, and this inhibition is relieved in cells treatedwith Shh. A Smo agonist stimulated and a Smo antagonist (cyclopamine)inhibited both phosphorylation of Smo by GRK2 and interactionof ß-arrestin 2 with Smo. ß-Arrestin 2 andGRK2 are thus potential mediators of signaling by activatedSmo.
1 Department of Medicine, Duke University Medical Center, Durham, NC 27710, USA. 2 Howard Hughes Medical Institute, Departments of Medicine and Biochemistry, Duke University Medical Center, Durham, NC 27710, USA. 3 Department of Cell Biology, Duke University Medical Center, Durham, NC 27710, USA. 4 Howard Hughes Medical Institute, Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA. 5 Department of Molecular Oncology, Genentech, South San Francisco, CA 94080, USA.
Present address: Department of Molecular Pharmacology, StanfordUniversity School of Medicine, Stanford, CA 94305, USA
* To whom correspondence should be addressed. E-mail: lefko001{at}receptor-biol.duke.edu (R.J.L.) and w.chen{at}duke.edu (W.C.).
Smoothened Signaling in Vertebrates Is Facilitated by a G Protein-coupled Receptor Kinase.
M. Philipp, G. B. Fralish, A. R. Meloni, W. Chen, A. W. MacInnes, L. S. Barak, and M. G. Caron (2008)
Mol. Biol. Cell
19, 5478-5489
|Abstract »|Full Text »|PDF »
{beta}-Arrestin-Mediated Localization of Smoothened to the Primary Cilium.
J. J. Kovacs, E. J. Whalen, R. Liu, K. Xiao, J. Kim, M. Chen, J. Wang, W. Chen, and R. J. Lefkowitz (2008)
Science
320, 1777-1781
|Abstract »|Full Text »|PDF »
A Kinetic Model for Calcium Dynamics in RAW 264.7 Cells: 1. Mechanisms, Parameters, and Subpopulational Variability.
The G protein-coupled receptor regulatory kinase GPRK2 participates in Hedgehog signaling in Drosophila.
C. Molnar, H. Holguin, F. Mayor Jr., A. Ruiz-Gomez, and J. F. de Celis (2007)
PNAS
104, 7963-7968
|Abstract »|Full Text »|PDF »
Immunosuppressive and Anti-angiogenic Sphingosine 1-Phosphate Receptor-1 Agonists Induce Ubiquitinylation and Proteasomal Degradation of the Receptor.
M. L. Oo, S. Thangada, M.-T. Wu, C. H. Liu, T. L. Macdonald, K. R. Lynch, C.-Y. Lin, and T. Hla (2007)
J. Biol. Chem.
282, 9082-9089
|Abstract »|Full Text »|PDF »
Analysis of Nsdhl-deficient embryos reveals a role for Hedgehog signaling in early placental development.
Arrestin-2 and G Protein-coupled Receptor Kinase 5 Interact with NF{kappa}B1 p105 and Negatively Regulate Lipopolysaccharide-stimulated ERK1/2 Activation in Macrophages.
N. Parameswaran, C. S. Pao, K. S. Leonhard, D. S. Kang, M. Kratz, S. C. Ley, and J. L. Benovic (2006)
J. Biol. Chem.
281, 34159-34170
|Abstract »|Full Text »|PDF »
Cardiac-Specific Ablation of G-Protein Receptor Kinase 2 Redefines Its Roles in Heart Development and {beta}-Adrenergic Signaling.
S. J. Matkovich, A. Diwan, J. L. Klanke, D. J. Hammer, Y. Marreez, A. M. Odley, E. W. Brunskill, W. J. Koch, R. J. Schwartz, and G. W. Dorn II (2006)
Circ. Res.
99, 996-1003
|Abstract »|Full Text »|PDF »
Smoothened Signal Transduction Is Promoted by G Protein-Coupled Receptor Kinase 2.
A. R. Meloni, G. B. Fralish, P. Kelly, A. Salahpour, J. K. Chen, R. J. Wechsler-Reya, R. J. Lefkowitz, and M. G. Caron (2006)
Mol. Cell. Biol.
26, 7550-7560
|Abstract »|Full Text »|PDF »
Human Receptors Patched and Smoothened Partially Transduce Hedgehog Signal When Expressed in Drosophila Cells.
M. De Rivoyre, L. Ruel, M. Varjosalo, A. Loubat, M. Bidet, P. Therond, and I. Mus-Veteau (2006)
J. Biol. Chem.
281, 28584-28595
|Abstract »|Full Text »|PDF »
Activation of heterotrimeric G proteins by Smoothened.
N. A. Riobo, B. Saucy, C. DiLizio, and D. R. Manning (2006)
PNAS
103, 12607-12612
|Abstract »|Full Text »|PDF »
The Contributions of Protein Kinase A and Smoothened Phosphorylation to Hedgehog Signal Transduction in Drosophila melanogaster.
Arrestin-related proteins mediate pH signaling in fungi.
S. Herranz, J. M. Rodriguez, H.-J. Bussink, J. C. Sanchez-Ferrero, H. N. Arst Jr., M. A. Penalva, and O. Vincent (2005)
PNAS
102, 12141-12146
|Abstract »|Full Text »|PDF »
Loss of the Serine/Threonine Kinase Fused Results in Postnatal Growth Defects and Lethality Due to Progressive Hydrocephalus.
M. Merchant, M. Evangelista, S.-M. Luoh, G. D. Frantz, S. Chalasani, R. A. D. Carano, M. van Hoy, J. Ramirez, A. K. Ogasawara, L. M. McFarland, et al. (2005)
Mol. Cell. Biol.
25, 7054-7068
|Abstract »|Full Text »|PDF »
Dynamic Interaction between the Dual Specificity Phosphatase MKP7 and the JNK3 Scaffold Protein {beta}-Arrestin 2.
Present address: Department of Molecular Pharmacology, Stanford University School of Medicine, Stanford, CA 94305, USA 
