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Cardiac-specific deletion of voltage dependent anion channel 2 leads to dilated cardiomyopathy by altering calcium homeostasis

ABSTRACT: Voltage dependent anion channel 2 (VDAC2) is an outer mitochondrial membrane porin known to play a significant role in apoptosis and calcium signaling. Abnormalities in calcium homeostasis often leads to electrical and contractile dysfunction and can cause dilated cardiomyopathy and heart failure. However, the specific role of VDAC2 in intracellular calcium dynamics and cardiac function is not well understood. To elucidate the role of VDAC2 in calcium homeostasis, we generated a cardiac ventricular myocyte-specific developmental deletion of Vdac2 in mice. Our results indicate that loss of VDAC2 in the myocardium causes severe impairment in excitation-contraction coupling by altering both intracellular and mitochondrial calcium signaling. We also observed adverse cardiac remodeling which progressed to severe cardiomyopathy and death. Reintroduction of VDAC2 in 6-week-old knock-out mice partially rescued the cardiomyopathy phenotype. Activation of VDAC2 by efsevin increased cardiac contractile force in a mouse model of pressure-overload induced heart failure. In conclusion, our findings demonstrate that VDAC2 plays a crucial role in cardiac function by influencing cellular calcium signaling. Through this novel role in cellular calcium dynamics and excitation-contraction coupling VDAC2 emerges as a plausible therapeutic target for heart failure.

ORGANISM(S): Mus musculus

PROVIDER: GSE168487 | GEO | 2021/03/09

REPOSITORIES: GEO

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Project description:Background: Modulation of mRNA splicing acts as an important layer of gene regulation, in addition to transcriptional regulation and epigenetic modifications. RNA binding proteins (RBPs) play essential roles in mediating RNA splicing and are key regulators of heart development and function. Our previous studies demonstrated that RBPMS (RNA-binding protein with multiple splicing) regulates cardiac development through modulating mRNA splicing during embryogenesis. Here we explored the postnatal function of RBPMS in the heart. Methods: We ablated Rbpms in the heart by generating a cardiac-specific knockout mouse line (Myh6-Cre, Rbpmsfl/fl), and evaluated its cardiac functions by histology, echocardiography, and gene expression. Paired-end RNA sequencing and RT-PCR were performed to identify and validate splicing targets of RBPMS in adult mouse hearts. Proximity-dependent Biotin Identification (BioID) assay and mass spectrometry analysis were performed to identify RBPMS binding partners. We also measured contractility and calcium fluxes in isolated mouse cardiomyocytes, and contractile forces of cardiac papillary muscle. Human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CMs) were also used as a model to explore the influence of RBPMS on contractility of human cardiomyocytes. Results: he absence of Rbpms in the heart led to dilated cardiomyopathy (DCM) and heart failure, causing early death in mice. Mice with cardiac-specific knockout of Rbpms showed myocardium noncompaction with reduced cardiomyocyte number at the neonatal stage and developed DCM with pervasive myocardial fibrosis in adulthood. We found that RBPMS mediates a largely distinct RNA splicing profile in adult mouse hearts compared to neonatal hearts, indicating a stage-specific modulation of alternative RNA splicing by RBPMS. In adult hearts, RBPMS mainly influenced alternative splicing of genes associated with sarcomere structures and cardiomyocyte contraction, such as Ttn, Pdlim5 and Nexn, to generate new protein isoforms. In neonatal hearts, RBPMS influenced the splicing of cytoskeletal genes. RBMPS was associated with spliceosome factors and other RNA binding proteins that play important roles in the heart, such as RBM20 and GATA4. Importantly, we found that the absence of Rbpms caused severe cardiomyocyte contractile defects and reduced calcium sensitivity in both mouse and hiPSC-CMs. Our results demonstrated that Rbpms is crucial for postnatal cardiac function and cardiomyocyte contractility by regulating RNA alternative splicing. Conclusions: Loss of Rbpms in the heart causes reduced cardiomyocyte number and impaired cardiomyocyte contraction, leading to DCM and heart failure.

Project description:BACKGROUND: Cardiac hypertrophy compensates for increased biomechanical stress of the heart induced by prevalent cardiovascular pathologies, but can result in cardiac failure if left untreated. We hypothesized that the tail-anchored protein Dysferlin with multiple Ca2+-binding C2-domains is critical for the integrity of the transverse-axial tubule (TAT) network inside cardiomyocytes, and contributes to the proliferation of TAT endomembranes during pressure overload-induced cardiac hypertrophy. OBJECTIVE: To reveal the impact of the membrane fusion and repair protein Dysferlin on TAT network stabilization and proliferation necessary for the hypertrophic growth of cardiomyocytes. METHODS AND RESULTS: Super-resolution light and electron microscopy of mouse cardiomyocytes identified a specific localization of Dysferlin in a vesicular compartment in nanometric proximity to contact sites of the TAT network with the sarcoplasmic reticulum (SR), a.k.a. junctional complexes for Ca2+-induced Ca2+ release. Mass spectrometry was used to characterize the cardiac Dysferlin interactome, thereby identifying a novel protein interaction with the membrane-tethering SR protein Juncophilin-2, a known interactor of L-type Ca2+ channels and Ryanodine Receptor Ca2+ release channels in junctional complexes. While the Dysferlin knockout caused a mild progressive phenotype of dilated cardiomyopathy in the mouse heart, global proteome analysis revealed changes preceding systolic failure. Following transverse aortic constriction (TAC), Dysferlin protein expression was significantly increased in hypertrophied wild-type myocardium, while Dysferlin knockout animals presented markedly reduced left-ventricular hypertrophy. Live-cell membrane imaging demonstrated a profound reorganization of the TAT network in wild-type left-ventricular myocytes post-TAC with robust proliferation of axial tubules, which critically depended on the increased expression of Dysferlin within newly-emerging tubule components. CONCLUSIONS: Dysferlin represents a new molecular target in cardiac disease that protects the integrity of tubule-SR junctional complexes for regulated excitation-contraction coupling, and controls TAT network reorganization and tubular membrane proliferation in cardiomyocyte hypertrophy induced by pressure-overload.

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Cardiac-specific deletion of voltage dependent anion channel 2 leads to dilated cardiomyopathy by altering calcium homeostasis

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