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.
Chen
Zhangguo
Z
Immunology Division, Department of Pediatrics, Faculty of Medicine, Université de Sherbrooke, 3001 N. 12th Avenue, Sherbrooke, QC J1H 5N4, Canada.
Gaudreau
Rémi
R
Le Gouill
Christian
C
Rola-Pleszczynski
Marek
M
Stanková
Jana
J
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