Fig. 3: Effects of ceramide application on conduction and gating properties of hERG1 variants expressed in mammalian HEK cells.
a Dose-response relationship for increasing [CER6] = 2, 20, 100, and 500 µM. Data are represented as mean ± SD. Raw values for residual current are provided in Source Data—Fig. 3a. Panels b and c show the contact mapping for ceramides calculated from CG-MD simulations for hERG1A-WT and hERG1A-F656C/F557L open-state (n = 4 for both), respectively. The extracellular view of the channel is shown, where the VSDs and the PD are shown as green and blue cartoons, respectively. The residues identified using the maximum-occupancy criteria are represented with a red surface. The residues identified through contact-duration but not through maximum-occupancy metrics are highlighted as a yellow surface. A color-coded list of residues is provided in Source Data—Fig. 3b, c. For each system, the contact-duration and maximum-occupancy metrics were calculated considering the tetrameric structure of the channel, totalling 4 × n replicas per-residue (see Source Data—Supplementary Figs. 5 and 6). The last 8 microseconds of each replica were used for analysis. Panels d and e show scatter plots for pIC50 (−log[IC50]) vs the change (Δ = Control − Cer) in deactivation τ and activation V0.5 after ceramide application, respectively (see Source Data—Supplementary Fig. 2b, c). The scatter plots for pIC50 vs τ and activation V0.5 after ceramide application are shown in Supplementary Fig. 13a, b, respectively. Deactivation τ values in d were measured at −100 mV, except for hERG1B (−50 mV). [CER6] = 20 µM was used for all measurements shown in panels d and e, except for the mutant hERG1A-F656C (400 µM). The legend for scatter plots for d and e is shown at the bottom: hERG1A- WT (orange), Y652A (yellow), M651T (green), F557L (blue), F656C (purple), and hERG1B (gray). The tables in a and d, e indicate the number of independent experiments (n), each performed on a different cell.
