Fig. 6: Ib30 potentiates endosomal trafficking of βarr1 for the V₂R^T360A mutant.

a Schematic representation of BRET-based endosomal localization assay for βarr1. b Co-expression of Ib30 promotes endosomal trafficking of βarr1 for V₂R^T360A as assessed by BRET assay. HEK-293 cells expressing the indicated constructs were stimulated with varying doses of AVP followed by BRET measurement. Data (mean ± SEM) from three independent experiments are presented here (Two-way ANOVA, Tukey’s multiple comparisons test; ****p < 0.0001). c Comparison of ΔBRET (difference in the BRET signal at the lowest and highest dose of AVP) in the BRET assay based on the data presented in panel B from three independent experiments (mean ± SEM, One-way ANOVA, Tukey’s multiple comparisons test; **p < 0.01, ****p < 0.0001). d Schematic representation of NanoBiT-based endosomal localization assay for βarr1. e Co-expression of Ib30 robustly promotes endosomal trafficking of βarr1 for V₂R^T360A as assessed by NanoBiT assay. HEK-293 cells expressing the V₂R^WT or V₂R^T360A, together with SmBiT-tagged βarr1 and Ib-CTL/Ib30 were stimulated with indicated doses of AVP followed by luminescence measurement. Data (mean ± SEM) from five independent experiments, normalized with maximal response under V₂R^WT + Ib-CTL condition (treated as 100%) (Two-way ANOVA, Tukey’s multiple comparisons test; ****p < 0.0001) are presented here. f Comparison of maximal response (at 1 µM AVP) in the NanoBiT-based endosomal trafficking assay presented in panel E from five independent experiments (mean ± SEM, One-way ANOVA, Tukey’s multiple comparisons test; **p < 0.01, ***p < 0.001, ns = non-significant). Source data are provided as a Source Data file.