Fig. 1: Expression and activation of the HyPer-DAO fusion protein in cardiomyocyte-specific transgenic mice.

A Scheme of the HyPer-DAO construct used for generating HyPer-DAO cardiomyocyte-specific transgenic mice and the HyPer and DAO chemical reactions. B Immunoblot for HyPer-DAO and GAPDH protein levels in cardiac protein lysates from three independent wild type (wt) and three independent HyPer-DAO mice. C Cardiac-specific HyPer-DAO protein expression in a HyPer-DAO mouse as demonstrated by immunoblot performed with protein lysates from different organs. This confirmatory experiment has been performed once. D Representative confocal image of a DRAQ5 stained cardiomyocyte isolated from a HyPer-DAO mouse. In total cardiomyocytes from two independent mice were isolated and 5 cardiomyocytes per mouse were imaged. E Cardiac output (CO) as determined by echocardiography in resting male and female HyPer-DAO and wt mice (n = independent 13 wt male, 15 HyPer-DAO male, 16 wt female and 20 HyPer-DAO female mice). F Response of HyPer-DAO cardiomyocytes to treatment with D-ala or L-ala (added at time point 4 min). The HyPer fluorescence response was recorded in the cytosol (upper panel) and the nucleus (lower panel). Ratios are normalized to the HyPer ratio prior to treatment, n = 27, 32, 47, 56, 38 and 13 independent cardiomyocytes were analyzed regarding their response in the nucleus and n = 23, 26, 13, 29, 25, and 17 cardiomyocytes regarding their response in the cytoplasm were analyzed after treatment with 3 mM D-ala, 4 mM D-ala, 6 mM D-ala, 8 mM D-ala, 10 mM D-ala and 10 mM L-ala, respectively. scale bar, 10 µm. Data are presented as mean values ± SEM. Source data are provided as a Source Data file.