Fig. 3
Exon 8 deletion results in KATP channel loss-of-function. a KATP channel structure showing pore-forming Kir6.x subunits (blue) associated with two of four SUR subunits (gray) (PDB 5WUA [doi 10.2210/pdb5WUA/pdb]), with the equivalent position of the SUR2 amino acids encoded by ABCC9 exon 8 in red (Ala389-Gln440). b Western blot of whole-cell lysate from Cosm6 cells transiently transfected with GFP or Kir6.2 alongside either wild-type SUR2A-Flag (WT) or SUR2A-Flag∆8. Left: Immunoblots using a primary antibody targeting the Flag-tag of SUR2A-Flag∆8 (top) and actin control (bottom). Right: Normalized expression for actin and SUR2A-Flag from cells transfected with WT SUR2A-Flag or SUR2A-Flag∆8. The data from four independent experiments, * denotes p < 0.05 according to Mann–Whitney U Test. A representative example blot is included in the associated Source Data File. c Left: 86Rb+ efflux from Cosm6 cells transfected with GFP alone, Kir6.2 alongside WT SUR2A, SUR2A∆8, or a 1:1 ratio of WT SUR2A and SUR2A∆8 (WT:∆8). Inset: early time efflux/time data points used to derive efflux rate constants. Right: Efflux rate constants for cells transfected with WT SUR2A, SUR2A∆8, and 1:1 ratio of WT:∆8. The data from six replicates in three independent experiments, ****p < 0.0001 (Mann–Whitney U Test). d Left: 86Rb+ efflux experiment from Cosm6 cells transfected with GFP alone, with Kir6.2 alongside WT SUR2B or SUR2B∆8. Right: Efflux rate constants for cells transfected with WT SUR2B or SUR2B∆8. The data from six replicates in three independent experiments, ****p < 0.0001 (Mann–Whitney U Test). e Example inside–out patch clamp recordings from Cosm6 cells transfected with Kir6.2 alongside SUR2A-WT (black), SUR2A∆8 (red), or a 1:1 mix of SUR2A-WT and SUR2A∆8 to mimic heterozygous expression (blue). The membrane potential was held at −50 mV in symmetrical KINT solutions, and ATP was applied as indicated. Arrows indicate the point of patch excision. f KATP currents from excised patches. g ATP dose–response relationship for SUR2A-WT or 1:1 SUR2A-WT:SUR2A∆8 channels. Inset: summary of ATP IC50 values. The data from individual experiments shown as dots alongside mean ± SEM. Source data are provided as a Source Data file
