Figure 3.

Heterodimerization of NR2E3 with retinal transcription factors. A) HEK293T cells were transfected with 3 μg of RLuc-NR2E3 wild-type (RLucNR2E3 or RN) and with 3 μg of GFP-NR2E3 wild-type (GFPNR2E3), GFP-TLX/NR2E1 (GFPTLX), GFP-RXRα/NR2C1 (GFPRXR), GFP-CRX (GFPCRX), GFP-NRL (GFPNRL) and GFP- rev-erbα/NR1D1 (GFP-NR1D1) expression plasmids. Experiments were performed four times in duplicates and SEM indicated. For statistical analysis, one-way ANOVA followed by Dunnett’s multiple comparison test comparing the mean of each column with the mean of the control RLucNR2E3 column was performed. B–D) Differential interaction of NR2E3 variant proteins with CRX, NRL and NR1D1. HEK293T cells were transiently transfected with 3 μg of RLuc-CRX (A), RLuc-NRL (B) and RLuc-rev-erbα/NR1D1 (RLuc-NR1D1) (C) expression plasmids, and, 3 μg of GFPC3-NR2E3 wild-type (NR2E3WT) or NR2E3 variant expression plasmids. RLuc expression vectors alone (respectively CRXctrl, NRLctrl and NR1D1ctrl) were used as negative controls, and Rluc-NR2E3WT/GFP-NR2E3WT transfected as positive controls (NR2E3ctrl). Experiments were performed three times in duplicates and SEM indicated. For statistical analysis, one-way ANOVA followed by Dunnett’s multiple comparison test, comparing the mean of each column with the mean of the control NR2E3 wild-type (NR2E3WT) column was performed. E–G) Characterization of CRX/NRL/NR2E3 interactions. HEK293T cells were transiently transfected with 2 μg of RLuc-CRX (E, F) and RLuc-NRL (G) expression plasmids, and increasing amounts of GFPC3-NRL (E), GFPC3-NR2E3 (F, G), in presence or not of 2 μg of pcDNA3.1/HisC-hNR2E3 (E), pMT-NRL (F), and pcDNA3.1/HisC-hCRX (G) expression plasmids. Experiments were performed three times in duplicates and SEM indicated. For statistical analysis, two-way ANOVA was performed. *: p<0.05; **<0.01; ***: p<0.001; ****: p<0.0001.