Skip to main content
. 2022 Apr 21;13:2185. doi: 10.1038/s41467-022-29902-4

Fig. 2. Characterization of hearts lacking CMYA5.

Fig. 2

a Genomic structure of Cmya5 wild-type and Δ alleles. Cas9-mediated deletion of 9281 bp of exon 2 causes a frameshift (fs) after the 55th amino acid residue. qRTPCR and genotyping amplicons are indicated. b PCR genotyping using WT (1 + 2) and Δ (1 + 3) primers. Representative of five independent experiments. c Cmya5 cardiac mRNA levels. RT-qPCR amplicon 1 and 2 represent the deleted region and the 3’ end of the transcript, respectively. ANOVA with Dunnett’s multiple comparison test vs. WT of the same amplicon. n, number of hearts. Data are presented as mean ± SD. d Cardiac protein lysates were analyzed by western blotting. KO samples lacked CMYA5 immunoreactivity. Representative of five independent experiments. e Gross morphology of WT and KO hearts. Bar = 2.5 mm. Representative of three independent experiments. f Heart weight normalized to body weight, at the indicated ages. ANOVA with Dunnett’s multiple comparison test vs. WT at the same time point. n, number of hearts. g Echocardiographic measurement of systolic heart function. FS fractional shortening. n, sample size. Kruskal–Wallis with Dunn’s multiple comparison test vs. WT at the same time point. h In situ T-tubule imaging. After plasma membrane labeling by MM4-64. 3-month-old hearts were optically sectioned using a confocal microscope. Right, transverse, and longitudinal T-tubule fractions. Mann–Whitney. Bar = 10 µm. n, number of cells. i Isolated ventricular WT or KO cardiomyocytes immunostained for T-tubule (TT: CAV3) and jSR (RYR2, CASQ2, JPH2) markers, as well as FSD2. Bar = 10 μm. j–l Transmission electron microscopy of WT or KO ventricular myocardium. *, T-tubule. The WT T-tubule micrograph is enlarged and labeled in k. l Quantification of T-tubule parameters, defined in Supplementary Fig. 4a. n, number of dyads from at least 10 cardiomyocytes from 3 different mice. Mann–Whitney. Bar = 500 nm. m Isolated WT or KO atrial cardiomyocytes immunostained for RYR2 or FSD2. NS, not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001. Statistical tests were two-sided. Data are presented as mean ± SD.