Fig. 4: Designs of eGAL^+/+-based biosensors for reporting endogenous pathways.

a Schematic of eGAL^+/+-based GPCR signaling. Upon ligand stimulation, β-arrestin2-nTEVp-eGAL^+/+ would be recruited to AVPR2-cTEVp to reconstitute TEVp for the proteolytic release of membraned anchored eGAL^+/+. b Fluorescence images of vasopressin-induced reporter expression. Merged images of BFP (cyan, BFP plasmid was co-transfected as a control), GFP (green), mCherry (red), and bright field are shown. Scale bar = 50 μm. c Vasopressin-induced fold changes using eGAL^+/+ or GAL4 to activate reporter fluorescence after 24 h. Fold changes of mCherry fluorescence are labeled for each comparison. Dose-dependent profiles of (d) agonist vasopressin-induced and (e) antagonist VPA-985-inhibited β-arrestin2 recruitment. For inhibition, 5 nM of Vasopressin was added together with diffirent concentrations of VPA-985. f Schematic of eGAL^+/+-based sensing of cytosolic PPARγ/RXRα dimerization. Dose-response curves of (g) agonist-induced dimerization and (h) antagonist-inhibited dimerization of eGAL^+/+-nTEVp-PPARγ and cTEVp-RXRα. GW9662 was added together with 1 μM GW1929 or 15 μM Rosiglitazone. Error bars denote MMFI (arbs.units) ± SD, n = 3, individual data points represent independent triplicates performed on the same day. Significance (two-tailed, unpaired t test) is labeled above data, ns: not significant, *p < 0.05, ****p < 0.0001. a, f Created in BioRender. He, J. (2025) https://BioRender.com/y0i9dcq. Source data are provided as a Source Data file.