The association of arrestin-3 with the follitropin receptor depends on receptor activation and phosphorylation.
We have recently shown that the binding of arrestin-3 to the lutropin receptor (LHR) is dependent mostly on receptor activation rather than on phosphorylation. The experiments presented here were designed to test the involvement of these two events in the association of arrestin-3 with the closely related follitropin receptor (FSHR). Activation of the FSHR leads to the phosphorylation of residues in the first and third intracellular loops. Mutation of the phosphorylation sites in the third intracellular loop of the rat (r) FSHR partially reduces phosphorylation but has no effect on arrestin-3 association. Mutation of the phosphorylation sites in the first intracellular loop abolishes phosphorylation and arrestin-3 association. Dominant-negative mutants of G protein-coupled receptor kinase (GRKs) 2 and 6 inhibit rFSHR phosphorylation to the same extent but only the dominant-negative mutant of GRK2 inhibits arrestin-3 association. Two mutations of the rFSHR (D389N and Y530F) that impair activation and abolish phosphorylation also impair arrestin-3 binding. GRK2 restores the phosphorylation of both mutants but it restores arrestin-3 association only to the D389N mutant. We conclude that, in contrast to the data obtained with the LHR, the association of arrestin-3 with the FSHR is dependent on receptor phosphorylation. The phosphorylation of the third intracellular loop residues is not needed for arrestin-3 association, however.
Krishnamurthy
Hanumanthappa
H
Department of Pharmacology, The University of Iowa College of Medicine, 2-319A BSB, 51 Newton Road, Iowa City, IA 52242-1109, USA.
Galet
Colette
C
Ascoli
Mario
M
D019390:Arrestins / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000502:physiology; D054768:G-Protein-Coupled Receptor Kinases; D006801:Humans; D008856:Microscopy, Fluorescence; D009154:Mutation; D010766:Phosphorylation; D011485:Protein Binding / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000502:physiology; D011962:Receptors, FSH / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D051552:beta-Adrenergic Receptor Kinases
D019390:Arrestins; D011962:Receptors, FSH; C082333:arrestin3; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D054768:G-Protein-Coupled Receptor Kinases; C089176:G-protein-coupled receptor kinase 6