Fig. 5

Identification of host P2X1 purinergic receptor as a TgROP18 target protein by FRET and co-IP. a Representative confocal FRET images. COS7 cells were cultured in plates the day before transfection. The experimental group was co-transfected with pECFP-N1-ROP18 and pEYFP-C1-P2X1, the negative control with pECFP-N1 and pEYFP-C1, and the positive control was transfected with pEYFP-CFP. The cells were fixed for confocal FRET measurement at 48 h post-infection. The vertical color bars represent FRET intensity, with red for a high FRET signal and blue for a low signal. b Quantitative analysis of FRET efficiency. c Co-IP analysis of COS7 cells transfected with pcDNA3.1-ROP18-3×Flag and/or pcDNA3.1-P2X1-HA. Lysates of the COS7 cells transiently transfected with the indicated plasmids or the control cells were immunoprecipitated with the anti-HA antibody and detected by western blotting with the indicated antibodies. d Co-IP analysis of SF268 cells infected with RH-ROP18-GFP-Flag. Lysates of the infected SF268 cells or the control cells were immunoprecipitated with anti-Flag antibody and detected by western blotting with the indicated antibodies. e Co-IP analysis of COS7 cells transfected with pcDNA3.1-ROP18-3×Flag and/or pcDNA3.1-P2X1-HA, pcDNA3.1-P2X1Δ339-399-HA. Lysates of the transfected COS7 cells or the control cells were immunoprecipitated with the anti-HA antibody and detected by western blotting with the indicated antibodies. The FRET efficiency was evaluated from 4 areas. The values were analyzed using the Kruskal–Wallis H-test and Bonferroni correction (*P < 0.05, **P < 0.01)