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Fig. 6 | BMC Biology

Fig. 6

From: Cross-communication between Gi and Gs in a G-protein-coupled receptor heterotetramer guided by a receptor C-terminal domain

Fig. 6

Effect of interference peptides on recruitment of β-arrestin-2. a, b Receptor agonist-induced β-arrestin-2 recruitment was measured by BRET. HEK-293 T cells were transfected with the cDNAs for β-arrestin-2-Rluc (Arr-Rluc, 0.5 μg cDNA) and either A2A-YFP (0.4 μg cDNA) and A1R (0.4 μg cDNA) (a) or A1-YFP (0.4 μg cDNA) and A2AR (0.4 μg cDNA) (b). Cells were untreated (control) or treated for 4 h with 4 μM A2AR TAT-TM synthetic peptides (TM4–7, see Methods) before addition of medium (basal, gray bars) or 100 nM of either the A1R agonist CPA (black bars), the A2AR agonist CGS-21680 (CGS, white bars), or both (striped bars). Positive BRET was expressed as milli-BRET units (see Methods). Mean ± SEM (7 experiments/condition). One-way ANOVA followed by Bonferroni’s post-hoc test showed a significant effect over basal in samples treated with CGS-21680 or over forskolin in samples treated with CPA (*P < 0.05, **P < 0.01, ***P < 0.001). One-way ANOVA followed by Bonferroni’s post-hoc test showed a significant effect of CPA + CGS-21680 over CGS-21680 treatments (&P < 0.05, &&P < 0.01). c Intermolecular distances between the center of masses of the N- and C-domains of the A1R-bound arrestin and of the A2AR-bound arrestin obtained from molecular dynamics (MD) simulations of A1-A2AHet in complex with two molecules of β-arrestin-2. d Intermolecular distances between the center of mass of the N- and C-domains of the A2AR-bound arrestin and the Cα atom of Glu238 (RAS domain) of Gi obtained from MD simulations of A1-A2AHet in complex with Gi bound to A1R and β-arrestin-2 bound to A2AR. These intermolecular distances are depicted as double arrows in the adjacent representative snapshots of the molecular models. Arrestin is shown in gray, whereas the color code of the depicted proteins is as in Fig. 3

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