Fig. 3: Designs of eGAL+/+-based CID reporting systems. | Nature Communications

Fig. 3: Designs of eGAL+/+-based CID reporting systems.

From: A generalizable approach for programming protease-responsive conformationally inhibited artificial transcriptional factors

Fig. 3

a Schematic of CID-mediated reconstitution of the split TEVp by dimerization between FRB-cTEVp and FKBP-nTEVp-eGAL+/+ for the proteolytic activation of eGAL+/+. Dose-response curves of (b) RAP-induced dimerization of FRB-cTEVp and FKBP-nTEVp-eGAL+/+, and (c) ABA-induced dimerization of PYL1-cTEVp and ABI-nTEVp-eGAL+/+ for transcription activation of mCherry expression. d Schematic of comparison between eGAL+/+ and GAL4 in reporting the RAP-induced dimerization of membrane associated FKBP-nTEVp-(eGAL+/+ or GAL4) and FRB-cTEVp. Upon RAP induction, the dimerization of extracellular FKBP and FRB would reconstitute the intracellular TEVp for the proteolytic release of membraned anchored eGAL+/+ or GAL4. e Fluorescence images of reporter expression before and after RAP induction. 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. Dose-dependent profiles of RAP-induced reporter expression using (f) GAL4 or (g) eGAL+/+-based MESA. RAP-induced mCherry expression at 0, 12, 24, and 36 h post RAP incubation from (h) GAL4 or (i) eGAL+/+-based MESA. 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) and RAP-induced fold changes of mCherry fluorescence are labeled for each comparison. ns: not significant, *p < 0.05, ** p < 0.01, ***p < 0.001. a, d Created in BioRender. He, J. (2025) https://BioRender.com/wxg3f87. Source data are provided as a Source Data file.

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