The m.3243A>G mutation in mitochondrial tRNA-Leu(UUR) is one of the most common pathogenic mitochondrial DNA mutations in humans. The clinical manifestations are highly heterogenous and the causes for the drastic clinical variability are unknown. Approximately one third of patients suffer from cardiac disease, which often increases mortality. Why only some patients develop cardiomyopathy is unknown. Here, we studied the molecular effects of a high m.3243A>G mutation load on cardiomyocyte functionality, using cells derived from induced pluripotent stem cells (iPSC-CM) of two different m.3243A>G patients, only one of them suffering from severe cardiomyopathy. While high mutation load impaired mitochondrial respiration in both patients’ iPSC-CMs, the downstream consequences varied. mtDNA mutant cells from a patient with no clinical heart disease showed increased glucose metabolism and retained cellular ATP levels, whereas cells from the cardiac disease patient showed reduced ATP levels. In this patient, the mutations also affected intracellular calcium signaling, while this was not true in the other patient’s cells. Our results reflect the clinical variability in mitochondrial disease patients and show that iPSC-CMs retain tissue specific features seen in patients.

Transcriptional coactivator PGC-1α is a main regulator of cardiac energy metabolism. In addition to canonical PGC-1α1, other PGC-1α isoforms have been found to exert specific biological functions in a variety of tissues. We investigated the expression patterns and the biological effects of the non-canonical isoforms in the heart. We used RNA sequencing data to identify the expression patterns of PGC-1α isoforms in the heart. To evaluate the biological effects of the alternative isoform expression, we generated a transgenic mouse with cardiac-specific overexpression of PGC-1α4 and analysed the cardiac phenotype with a wide spectrum of physiological and biophysical tools. Our results show that non-canonical isoforms are expressed in the heart, and that the main variant PGC-1α4 is induced by β-adrenergic signalling in adult cardiomyocytes. Cardiomyocyte specific PGC-1α4 overexpression in mice relieves the RE1-Silencing Transcription factor (REST)-mediated suppression of neuronal genes during foetal heart development. The resulting de-repression of REST target genes induces a cardiac phenotype with increased cellular energy consumption, resulting in postnatal dilated cardiomyopathy. These results propose a new concept for actions of the PGC-1α protein family where activation of the Pgc-1α gene, through its isoforms, induces a phenotype with concurrent supply and demand for cellular energy. These data highlight the biological roles of the different PGC-1α isoforms, which should be considered when future therapies are developed.

Hypertrophic cardiomyopathy (HCM) is often characterized by a complex landscape of secondary compensatory changes, making it difficult to distinguish primary events triggered by the underlying genetic variants. Here, using patient-derived iPSC-engineered heart tissues cultured in conditions minimizing external stimuli, we identify the initial phenotype of the HCM inducing variant MYBPC3-Q1061X. We demonstrate that initially MYBPC3 haploinsufficiency leads to robust, muscle length-dependent hypercontractility with emerging diastolic dysfunction at higher contraction rates. This phenotype is entirely decoupled from early transcriptional and metabolic stress responses, as evidenced by physiological phenotype and metabolite and cell type-specific transcriptional profiles. Mathematical modeling reveals that the hypercontractile phenotype can be translated to adult cardiac muscle with a small change in myosin availability, amplified by inherent cooperativity within the sarcomere that is instrumental in enhancing the force and the speed of the contraction cycle. Despite the apparent calcium sensitization, these sarcomeric changes do not affect excitation-contraction coupling or calcium buffering in this proximal stage. These results suggest that the primary biophysical defect in MYBPC3-haploinsufficiency manifests as a latent mechanical hypersensitivity that precedes secondary cellular remodeling and functional instability. This initial phenotype provides a window for preventive therapeutic intervention before the onset of permanent cellular remodeling in the heart.