Fig. 6: Gαs–β-arrestin1 (βarr1) interaction and its functional consequence in the cell.

a Schematic illustration of the PLA-based Gαs–βarr1 interaction monitoring system. b Representative PLA images showing the interaction between Gαs and βarr1 under the indicated conditions (NT = non-treated; +AVP = Arginine Vasopressin-treated). PLA puncta are shown in gray, and Hoechst-stained nuclei in cyan. Scale bars: 10 μm. c Quantification of PLA signals. Data represent pooled counts from 42 (NT) and 44 (AVP) randomly selected fields of view across 3 independent experiments. Bars indicate mean ± SEM. Scatter plots show individual data points. *P < 0.0001; statistical analysis performed using unpaired two-tailed Student’s t-test. The exact P-value is 0.000001191. d Schematic illustration of BRET-based βarr1 β-strand XX (βXX) release sensing system. e Basal BRET ratio in the absence or presence of Gαs is shown as violin plots with the median line (left panel); corresponding luminescence intensities are presented as mean ± SD with individual points from 27 wells per condition (right panel); unpaired two-tailed Student’s t-test, *P < 0.001. The exact P-values for basal BRET ratio and luminescence intensity are 3.486 × 10⁻¹⁴ and 0.3768, respectively. f BRET values measured 10 min after AVP (100 nM) in the absence or presence of Gαs, shown as violin plots with the median line; data are from 9 wells per condition; one-way ANOVA with Tukey’s post-hoc test, *P < 0.0001. Exact P-values are as follows: Gαs–/–AVP vs. Gαs–/+AVP, 3.259 × 10⁻⁹; Gαs–/–AVP vs. Gαs+/–AVP, 0.000015; Gαs–/+AVP vs. Gαs+/+AVP, 0.000131; Gαs+/–AVP vs. Gαs+/+AVP, 2.499 × 10⁻¹⁰. g Proposed model of Gαs–βarr1 interaction and its functional outcome.