Fig 2: Nuclear IDH-2 regulates chromatin accessibility.a–e Accessibility of promoter region of cardioprotective genes ranging from anti-apoptotic (a), antioxidants (b), contractility related (c), involved in calcium handling (d), and metabolic genes (e) as assessed by ATAC-qPCR. n = 3 P value** <0.01, P value* <0.05.

Fig 3: Nuclear IDH-2 mediated changes in chromatin accessibility correspond to gene expression changes.a–e mRNA levels of anti-apoptotic genes (a), contractility-related genes (b), metabolic genes (c), calcium handling (d), and antioxidant genes (e). n = 3 P value**** <0.0001, P value** <0.009, P value* <0.05.

Fig 4: Expression of nuclear TCA dehydrogenase maintains the contractility of iPSC-CMs.a, b Calcium handling as assessed by Fluo-4 staining in cells expressing nuclear IDH-2 in the presence of DRN. IDH-2 expression in the nucleus prior to DRN treatment maintains the rhythmic change in calcium (a, b). n = 3 P value ****<0.0001, *<0.05. c–h Representative graph for contraction and relaxation cycle of cardiomyocyte contractility (c), quantification of beat rate (d), contraction velocity (e), relaxation velocity (f), maximum contraction (g), and maximum relaxation (h) in cells expressing nuclear IDH-2 prior to DRN treatment compared to control cells treated with DRN. n = 3 P value ****<0.0001, ***<0.0005, **<0.005. *=0.0399, i Representative M-mode images of DRN- or PBS-treated mice. j Left ventricle ejection fraction percentage and k fractional shortening recorded from M-mode tracings in mice treated with DRN or PBS. n = 3 P value*** <0.001. l Representative M-mode images of DRN-treated mice injected with GFP- or NLS-IDH2-expressing lentivirus. m Left ventricle ejection fraction percentage and n fractional shortening recorded from M-mode tracings in mice expressing GFP or NLS-IDH-2 prior to DRN treatment. P value *<0.05. n = 5 (GFP, both male and female), n = 8 (NLS-IDH-2, both male and female).

Fig 5: Nuclear IDH-2 induces an anti-apoptotic effect.a a-KG levels measured in isolated nuclei, n = 3 p value* <0.0130. b Schematic of the proposed a-KG-AMPK-Sirtuin- pathway. c Western blots showing increased AMPK phosphorylation upon nuclear IDH-2 expression. ß-actin was used as a loading control. d Sirtuin activity represented as OD/min/mg. n = 3 P value** = 0.0036. e Schematic of inhibitory sites of compound C and sirtinol in the a-KG-AMPK-Sirtuin pathway. f, g Caspase 9 activity (f) n = 3 p value* <0.05, p value*** = 0.0002, p value **** <0.0001, and cell death (g) n = 4 p value*** <0.0005, p value** = 0.0085 upon inhibition of AMPK and sirtuin by compound C and sirtinol, respectively. h Reduced gene expression of anti-apoptotic protein BCL2 by AMPK and sirtuin inhibition in cells expressing nuclear IDH-2. n = 3 p value** <0.05, p value* <0.002. i, j Annexin V staining in iPSC-CM expressing nuclear IDH-2 and treated with succinate in the presence of DRN (i) or expressing IDH-2 mutants in the nuclei of cells treated with DRN (j). n = 3 P value**** <0.0001.