FIGURE 8.

Dose-dependent effects of TM peptides on both opsin and isoRho dimerization in live cells. Effects of increasing concentrations of TM peptides on theBRET signal. HEK-293 (mOpsin·Rluc and mOpsin·Venus) cells were either incubated with 9-cis-retinal resulting in a light-sensitive isoRho at the cell membrane (A, C, and E) or without 9-cis-retinal so that the receptor remained in its opsin, light-insensitive state (B, D, and F). The BRET signal was measured after cells were treated with TM peptides at concentrations ranging from 0.02 to 2 μm for 1 h at 37 °C. BRET signals derived from three independent experiments obtained for TM1, TM2, TM4, and TM5 were scored as described under “Materials and Methods,” and a dose-response curve was fitted by the Hill function in SigmaPlot software. Calculated half-maximal BRET signal inhibition values are presented as EC₅₀ values (A–D). E and F, overlays of ΔBRET determined in TM peptide-treated cells indicate the inhibitory efficacies of these peptides. TM3, TM6, and TM7 peptides and the scrambled TM4 peptide had no inhibitory effect on the BRET signal.