TY - JOUR A1 - Janz, Anna A1 - Walz, Katharina A1 - Cirnu, Alexandra A1 - Surjanto, Jessica A1 - Urlaub, Daniela A1 - Leskien, Miriam A1 - Kohlhaas, Michael A1 - Nickel, Alexander A1 - Brand, Theresa A1 - Nose, Naoko A1 - Wörsdörfer, Philipp A1 - Wagner, Nicole A1 - Higuchi, Takahiro A1 - Maack, Christoph A1 - Dudek, Jan A1 - Lorenz, Kristina A1 - Klopocki, Eva A1 - Ergün, Süleyman A1 - Duff, Henry J. A1 - Gerull, Brenda T1 - Mutations in DNAJC19 cause altered mitochondrial structure and increased mitochondrial respiration in human iPSC-derived cardiomyocytes JF - Molecular Metabolism N2 - Highlights • Loss of DNAJC19's DnaJ domain disrupts cardiac mitochondrial structure, leading to abnormal cristae formation in iPSC-CMs. • Impaired mitochondrial structures lead to an increased mitochondrial respiration, ROS and an elevated membrane potential. • Mutant iPSC-CMs show sarcomere dysfunction and a trend to more arrhythmias, resembling DCMA-associated cardiomyopathy. Background Dilated cardiomyopathy with ataxia (DCMA) is an autosomal recessive disorder arising from truncating mutations in DNAJC19, which encodes an inner mitochondrial membrane protein. Clinical features include an early onset, often life-threatening, cardiomyopathy associated with other metabolic features. Here, we aim to understand the metabolic and pathophysiological mechanisms of mutant DNAJC19 for the development of cardiomyopathy. Methods We generated induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) of two affected siblings with DCMA and a gene-edited truncation variant (tv) of DNAJC19 which all lack the conserved DnaJ interaction domain. The mutant iPSC-CMs and their respective control cells were subjected to various analyses, including assessments of morphology, metabolic function, and physiological consequences such as Ca\(^{2+}\) kinetics, contractility, and arrhythmic potential. Validation of respiration analysis was done in a gene-edited HeLa cell line (DNAJC19tv\(_{HeLa}\)). Results Structural analyses revealed mitochondrial fragmentation and abnormal cristae formation associated with an overall reduced mitochondrial protein expression in mutant iPSC-CMs. Morphological alterations were associated with higher oxygen consumption rates (OCRs) in all three mutant iPSC-CMs, indicating higher electron transport chain activity to meet cellular ATP demands. Additionally, increased extracellular acidification rates suggested an increase in overall metabolic flux, while radioactive tracer uptake studies revealed decreased fatty acid uptake and utilization of glucose. Mutant iPSC-CMs also showed increased reactive oxygen species (ROS) and an elevated mitochondrial membrane potential. Increased mitochondrial respiration with pyruvate and malate as substrates was observed in mutant DNAJC19tv HeLa cells in addition to an upregulation of respiratory chain complexes, while cellular ATP-levels remain the same. Moreover, mitochondrial alterations were associated with increased beating frequencies, elevated diastolic Ca\(^{2+}\) concentrations, reduced sarcomere shortening and an increased beat-to-beat rate variability in mutant cell lines in response to β-adrenergic stimulation. Conclusions Loss of the DnaJ domain disturbs cardiac mitochondrial structure with abnormal cristae formation and leads to mitochondrial dysfunction, suggesting that DNAJC19 plays an essential role in mitochondrial morphogenesis and biogenesis. Moreover, increased mitochondrial respiration, altered substrate utilization, increased ROS production and abnormal Ca\(^{2+}\) kinetics provide insights into the pathogenesis of DCMA-related cardiomyopathy. KW - cell biology KW - molecular biology KW - dilated cardiomyopathy with ataxia KW - genetics KW - metabolism KW - mitochondria KW - OXPHOS KW - ROS KW - contractility Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-350393 SN - 2212-8778 VL - 79 ER - TY - JOUR A1 - Ohlebusch, Barbara A1 - Borst, Angela A1 - Frankenbach, Tina A1 - Klopocki, Eva A1 - Jakob, Franz A1 - Liedtke, Daniel A1 - Graser, Stephanie T1 - Investigation of alpl expression and Tnap-activity in zebrafish implies conserved functions during skeletal and neuronal development JF - Scientific Reports N2 - Hypophosphatasia (HPP) is a rare genetic disease with diverse symptoms and a heterogeneous severity of onset with underlying mutations in the ALPL gene encoding the ectoenzyme Tissue-nonspecific alkaline phosphatase (TNAP). Considering the establishment of zebrafish (Danio rerio) as a new model organism for HPP, the aim of the study was the spatial and temporal analysis of alpl expression in embryos and adult brains. Additionally, we determined functional consequences of Tnap inhibition on neural and skeletal development in zebrafish. We show that expression of alpl is present during embryonic stages and in adult neuronal tissues. Analyses of enzyme function reveal zones of pronounced Tnap-activity within the telencephalon and the mesencephalon. Treatment of zebrafish embryos with chemical Tnap inhibitors followed by axonal and cartilage/mineralized tissue staining imply functional consequences of Tnap deficiency on neuronal and skeletal development. Based on the results from neuronal and skeletal tissue analyses, which demonstrate an evolutionary conserved role of this enzyme, we consider zebrafish as a promising species for modeling HPP in order to discover new potential therapy strategies in the long-term. KW - nonspecific alkaline-phosphae KW - in situ hybridization KW - hypophosphatasia KW - promotes KW - model KW - neurotransmission KW - differentiation KW - mineraliztion KW - metabolism KW - vertebrate Y1 - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-230024 VL - 10 ER -