Fig. 7: Genetic and pharmacologic rescue of NAA10-mutant iPSC-CMs.
From: Dysregulation of N-terminal acetylation causes cardiac arrhythmia and cardiomyopathy
a iPSC-CMs were transduced with adenoviruses encoding LacZ or NAA10-P2A-HaloTag. Created in BioRender. Yoshinaga, D. (2025) https://BioRender.com/p38t891. b Incubation with HaloTag dye indicates the expression of NAA10 (left lower) compared to LacZ (left upper). Scale bars: 100 μm. Halo-positive cells showed 84.3 ± 0.7 % of cells were transduced (n = 3) (right). c Western blot of whole-cell lysates from pNAA10R4S/Y- or eNAA10R4S/Y-transduced with Ad-LacZ or Ad-NAA10 and probed with anti-NAA10. The faint lower band represents native NAA10 expression (gray triangle). The intensity of NAA10 was quantified and normalized to vinculin (n = 1). MW: molecular weight. d Representative MEA recordings of iPSC-CM monolayers co-transduced with Ad-ChR2-GFP and either Ad-LacZ or Ad-NAA10, optically paced at 1 Hz (blue arrowheads). e Field potential duration (FPD) 48 hours post-transduction. NAA10 replacement significantly shortened FPD in pNAA10R4S/Y (299.8 ± 13.9 ms vs 255.4 ± 13.0 ms, p = 0.0015) and eNAA10R4S/Y (247.6 ± 6.4 vs 195.2 ± 5.7 ms, p = 0.0003), while it did not in WT (176.8 ± 13.0 ms vs 149.4 ± 10.3 ms, p = 0.0978). (three independent differentiation experiments; mixed-effects analysis). f MEA monolayers were measured at baseline and treated with the specific INaL blocker GS967. GS967 significantly shortened FPD in WT (167.7 ± 4.6 ms vs 154.1 ± 2.4 ms, p = 0.0379), pNAA10R4S/Y (352.2 ± 7.1 ms vs 272.5 ± 4.5 ms, p < 0.0001), and eNAA10R4S/Y (300.1 ± 6.6 ms vs 251.5 ± 5.6 ms, p < 0.0001). (three independent differentiation experiments; p = 0.0001; mixed-effects analysis). g Micro-patterned iPSC-CMs transduced with Ad-NAA10 were stained for sarcomeric alpha-actinin (SAA) to quantify changes in sarcomeric organization (scale bar: 10 μm). Automated analysis of single iPSC-CMs on micro-patterned substrates for h. sarcomeric packing density (Ad-NAA10(-) = 0.30 ± 0.01 vs Ad-NAA10(+) = 0.35 ± 0.02; three independent differentiation; p = 0.0734; two-tailed unpaired t-test) and i. Sarcomere length (Ad-NAA10(-) = 1.83 ± 0.03 vs Ad-NAA10(+) = 1.67 ± 0.04; three independent differentiation; p = 0.0063; two-tailed unpaired t-test). Data are presented as mean ± SEM. ****p < 0.0001, ***p < 0.001, **p < 0.01, ns = p > 0.1. Source data are provided as a Source Data file.
