Activator of G-protein signaling 3 (AGS3) binds GRK6 via its G-protein (GPR) domain to modulate CXCR2 mediated cellular responses. (CCR5P.261)

Abstract

Abstract AGS3, a receptor-independent activator of G-protein signaling, comprises of seven tetratricopeptide (TPR) motifs and four GPR motifs joined by a linker sequence. Previous studies have demonstrated that TPR domain of AGS3 is important for determining the positioning of AGS3 within the cell, whereas, GPR domain serves as a docking site for Gαi-GDP leading to AGS3-Gαi signaling. We have recently shown that GRK6 complexes with AGS3-Gαi to regulate CXCR2-mediated cellular functions including Ca2+ mobilization, mitogen-activated protein kinase (MAPK) activation, receptor desensitization and down-regulation. In the current study mutants from the GPR and TPR domains were used to determine which motif(s) of AGS3 is (are) involved in binding with GRK6. Co-imunoprecipitation studies in RBL cells co-transfected with wild type and mutants AGS3 showed that, upon stimulation with interleukin 8 (IL-8/CXCL8), GRK6 interacts with the GPR domain of AGS3. Overexpression of AGS3 and the GPRs motif (ΔTPR7-link), but not the TPRs motif (TPR), in RBL-CXCR2 inhibited CXCL8-induced intracellular Ca2+ mobilization and MAPK activation. Taken together, these results indicated that AGS3 binds to GRK6 via the GPR domain to modulate CXCR2-mediated signal transduction.