NHERF-1 and the cytoskeleton regulate the traffic and membrane dynamics of G protein-coupled receptors.
The sodium-hydrogen exchange regulatory factor 1 (NHERF-1/EBP50) interacts with the C terminus of several G protein-coupled receptors (GPCRs). We examined the role of NHERF-1 and the cytoskeleton on the distribution, dynamics, and trafficking of the beta(2)-adrenergic receptor (beta(2)AR; a type A receptor), the parathyroid hormone receptor (PTH1R; type B), and the calcium-sensing receptor (CaSR; type C) using fluorescence recovery after photobleaching, total internal reflection fluorescence, and image correlation spectroscopy. beta(2)AR bundles were observed only in cells that expressed NHERF-1, whereas the PTH1R was localized to bundles that parallel stress fibers independently of NHERF-1. The CaSR was never observed in bundles. NHERF-1 reduced the diffusion of the beta(2)AR and the PTH1R. The addition of ligand increased the diffusion coefficient and the mobile fraction of the PTH1R. Isoproterenol decreased the immobile fraction but did not affect the diffusion coefficient of the beta(2)AR. The diffusion of the CaSR was unaffected by NHERF-1 or the addition of calcium. NHERF-1 reduced the rate of ligand-induced internalization of the PTH1R. This phenomenon was accompanied by a reduction of the rate of arrestin binding to PTH1R in ligand-exposed cells. We conclude that some GPCRs, such as the beta(2)AR, are attached to the cytoskeleton primarily via the binding of NHERF-1. Others, such as the PTH1R, bind the cytoskeleton via several interacting proteins, one of which is NHERF-1. Finally, receptors such as the CaSR do not interact with the cytoskeleton in any significant manner. These interactions, or the lack thereof, govern the dynamics and trafficking of the receptor.
Wheeler
David
D
Department of Pharmacology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15261, USA.
Sneddon
W Bruce
WB
Wang
Bin
B
Friedman
Peter A
PA
Romero
Guillermo
G
D000199:Actins / Q000378:metabolism; D000818:Animals; D016466:CHO Cells; D002118:Calcium / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D003599:Cytoskeleton / Q000378:metabolism*; D004705:Endocytosis; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D018613:Microscopy, Confocal; D008954:Models, Biological; D010750:Phosphoproteins / Q000502:physiology*; D017434:Protein Structure, Tertiary; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000502:physiology*
D000199:Actins; D010750:Phosphoproteins; D043562:Receptors, G-Protein-Coupled; D017923:Sodium-Hydrogen Exchangers; C093376:sodium-hydrogen exchanger regulatory factor; D002118:Calcium