Fig. 3: IDH3γ is redox modified in the hearts of HyPer-DAO mice after activation of DAO in vivo.

A Scheme of the redox-proteomics analysis performed with heart samples obtained from n = 3 independent male HyPer-DAO and n = 3 independent wt mice that were treated for 7 days with 55 mM D-ala in the drinking water. B Venn-diagrams demonstrate elevated yield of cysteine-containing unique peptides and total number of identified peptide spectra matches (PSM) by redox enrichment. C Volcano plot of the peptides identified in the redox-proteomics screen as outlined in A. fold change (FC) D Protein dendrogram for the different murine isocitrate dehydrogenase isoforms and subunits. E Activity of the indicated TCA cycle enzymes determined in protein extracts obtained from hearts of 7 days D-ala treated female HyPer-DAO and wt mice. n = 6 independent mice per group. F Relative changes in RNA levels of IDH3 subunits in the hearts of HyPer-DAO mice versus wt mice after 7 days (n = 4 independent wt and n = 4 independent HyPer-DAO animals) and 21 days (n = 3 independent wt and n = 3 independent HyPer-DAO animals) of treatment with D-ala. G Immunoblot for IDH3γ and vinculin protein levels in heart samples of three independent wt and three independent HyPer-DAO mice after 7 days of treatment with D-ala. H IDH3 activity in samples obtained from hearts of a mixed group of male and female HyPer-DAO or wt mice after 7 days treatment with D-ala (n = 16 independent wt and n = 19 independent HyPer-DAO animals) or after 7 days treatment with D-ala plus 2 additional D-ala free days (off, n = 7 independent HyPer-DAO animals). I IDH1/2 and IDH3 activity in protein samples obtained from isolated HyPer-DAO cardiomyocytes non-treated (n.t.) and after treatment with 2 mM D-ala or L-ala for 20 min. Cardiomyocytes were isolated from n = 8 independent HyPer-DAO mice. J IDH3 activity determined in protein extracts obtained from the left ventricular myocardium of mice 4 weeks after transverse aortic constriction (TAC) versus sham surgery. n = 9 independent mice per group. K Representative immunoblot for IDH3γ oxidation analyzed by PEG switch assay in left ventricular samples from TAC and sham treated mice. The experiment has been performed three times. Data are presented as mean values ± SEM, two-tailed unpaired t-test (C, E and J), two-tailed one-sample t-test (F), one-way ANOVA (I) and one-way Welch’s ANOVA test (H). Source data are provided as a Source Data file.