Agonist-induced internalization of leukotriene B(4) receptor 1 requires G-protein-coupled receptor kinase 2 but not arrestins.
The leukotriene B(4) (LTB(4)) receptor (BLT1) becomes desensitized upon repeated agonist stimulation. Little is known, however, about BLT1 internalization, which follows desensitization in most G-protein-coupled receptors (GPCR). In the current study, transiently expressed BLT1 readily internalized, after LTB(4) stimulation, in RBL-2H3 cells that express high levels of endogenous GPCR kinase 2 (GRK2) but did not in COS-7 or human embryonic kidney (HEK) 293 cells, which do not overexpress GRK. The internalization of BLT1 could be blocked in RBL-2H3 cells by coexpressing dominant-negative (DN) GRK2 K220R and could be promoted in HEK293 cells by coexpressing wild-type (WT) GRK2. Coexpression of WT or DN nonvisual arrestins had no effect on BLT1 internalization. Moreover, upon stimulation with LTB(4), BLT1 did not induce arrestin-green fluorescence protein redistribution in either cell type, even in the presence of overexpressed GRK2. Coimmunoprecipitation experiments confirmed that BLT1 could associate with GRK2 but not with arrestins. A C-tail-truncated mutant of BLT1 lost the capacity to internalize and associate with GRK2 upon exposure to LTB(4), suggesting that the C-tail was required for receptor internalization and association with GRK2. Taken together, our results indicate that the C terminus of BLT1 plays a pivotal role in receptor internalization and GRK2 association. Moreover, ligand-induced BLT1 internalization is dependent on GRK2 but independent of arrestins. This may allow differential, cell-type-specific signaling in response to LTB(4), depending on GRK expression levels.
Immunology Division, Department of Pediatrics, Faculty of Medicine, Université de Sherbrooke, 3001 N. 12th Avenue, Sherbrooke, QC J1H 5N4, Canada.
Zhangguo
Z
Chen
Rémi
R
Gaudreau
Christian
C
Le Gouill
Marek
M
Rola-Pleszczynski
Jana
J
Stanková
D000818:Animals; D019390:Arrestins / Q000378:metabolism*; D019556:COS Cells; D002478:Cells, Cultured; D002966:Clathrin / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D034281:Dynamins / Q000378:metabolism; D004705:Endocytosis / Q000502:physiology; D006801:Humans; D007975:Leukotriene B4 / Q000378:metabolism; D018102:Receptors, Leukotriene B4 / Q000819:agonists*; D018048:Receptors, Purinergic P2 / Q000378:metabolism; D051552:beta-Adrenergic Receptor Kinases
D019390:Arrestins; D002966:Clathrin; C513240:LTB4R protein, human; D018102:Receptors, Leukotriene B4; D018048:Receptors, Purinergic P2; D007975:Leukotriene B4; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D034281:Dynamins