Fig. 2. Gq inhibition in Ang II mimics the GRK-subtype selectivity of TRV027.

a, Luminescent kinetics of β-arrestin1-recruitment responses upon 1 µM Ang II stimulation in the presence of pretreatment of the Gq inhibitor, YM-254890 (1 µM). b, Comparison of Ang II-induced β-arrestin1 recruitment in the parent and the GRK-deficient cell lines in the presence or absence of YM-254890. For each experiment, Ang II-induced β-arrestin1-recruitment signal in the parent cells was set as 100%. c, d, Assessment of the individual GRK subtypes for their sensitivity to YM-254890. The C-terminally FLAG epitope-tagged GRK2, 3, 5 and 6 constructs were individually expressed at a level equivalent to that of endogenous GRK2 (Supplementary Fig. 3a, b) along with AT1R-Sm and Lg-β-arrestin1 in ΔGRK2/3/5/6 cells and stimulated with 1 µM Ang II. For each experiment, Ang II-induced β-arrestin1-recruitment signal was set as 100% (d). Cell lines used were ∆GRK2/3, CL2; ∆GRK5/6, CL2; ∆GRK2/3/5/6, CL2 (a–d). In Fig. 2a, c, lines and shaded regions represent mean and SEM, respectively, of 3 independent experiments with each performed in duplicate. In Fig. 2b, d, bars and error bars represent mean and SEM, respectively, of 3 independent experiments with each performed in duplicate. In Fig. 2b, d, * and ** represent P < 0.05 and 0.01, respectively, with two-way ANOVA followed by the Dunnett’s test for multiple comparison analysis with reference to the Ang II stimulation. ns, not significantly different between the groups. See Supplementary statistics data file for additional statistics and exact P values.