TY - INPR A1 - Brenner, Marian A1 - Zink, Christoph A1 - Witzinger, Linda A1 - Keller, Angelika A1 - Hadamek, Kerstin A1 - Bothe, Sebastian A1 - Neuenschwander, Martin A1 - Villmann, Carmen A1 - von Kries, Jens Peter A1 - Schindelin, Hermann A1 - Jeanclos, Elisabeth A1 - Gohla, Antje T1 - 7,8-Dihydroxyflavone is a direct inhibitor of pyridoxal phosphatase T2 - eLife N2 - Vitamin B6 deficiency has been linked to cognitive impairment in human brain disorders for decades. Still, the molecular mechanisms linking vitamin B6 to these pathologies remain poorly understood, and whether vitamin B6 supplementation improves cognition is unclear as well. Pyridoxal phosphatase (PDXP), an enzyme that controls levels of pyridoxal 5’-phosphate (PLP), the co-enzymatically active form of vitamin B6, may represent an alternative therapeutic entry point into vitamin B6-associated pathologies. However, pharmacological PDXP inhibitors to test this concept are lacking. We now identify a PDXP and age-dependent decline of PLP levels in the murine hippocampus that provides a rationale for the development of PDXP inhibitors. Using a combination of small molecule screening, protein crystallography and biolayer interferometry, we discover and analyze 7,8-dihydroxyflavone (7,8-DHF) as a direct and potent PDXP inhibitor. 7,8-DHF binds and reversibly inhibits PDXP with low micromolar affinity and sub-micromolar potency. In mouse hippocampal neurons, 7,8-DHF increases PLP in a PDXP-dependent manner. These findings validate PDXP as a druggable target. Of note, 7,8-DHF is a well-studied molecule in brain disorder models, although its mechanism of action is actively debated. Our discovery of 7,8-DHF as a PDXP inhibitor offers novel mechanistic insights into the controversy surrounding 7,8-DHF-mediated effects in the brain. KW - 7,8-dihydroxyflavone (7,8-DHF) KW - pyridoxal phosphatase (PDXP) KW - vitamin B6 KW - PDXP inhibitors Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-350446 ER - TY - JOUR A1 - Eberl, Hanna A1 - Rebs, Sabine A1 - Hoppe, Stefanie A1 - Sedaghat-Hamedani, Farbod A1 - Kayvanpour, Elham A1 - Meder, Benjamin A1 - Streckfuss-Bömeke, Katrin T1 - Generation of an RBM20-mutation-associated left-ventricular non-compaction cardiomyopathy iPSC line (UMGi255-A) into a DCM genetic background to investigate monogenetic cardiomyopathies JF - Stem Cell Research N2 - RBM20 mutations account for 3 % of genetic cardiomypathies and manifest with high penetrance and arrhythmogenic effects. Numerous mutations in the conserved RS domain have been described as causing dilated cardiomyopathy (DCM), whereas a particular mutation (p.R634L) drives development of a different cardiac phenotype: left-ventricular non-compaction cardiomyopathy. We generated a mutation-induced pluripotent stem cell (iPSC) line in which the RBM20-LVNC mutation p.R634L was introduced into a DCM patient line with rescued RBM20-p.R634W mutation. These DCM-634L-iPSC can be differentiated into functional cardiomyocytes to test whether this RBM20 mutation induces development of the LVNC phenotype within the genetic context of a DCM patient. KW - cell biology KW - developmental biology KW - general medicine KW - RBM20 mutations KW - DCM genetic background KW - monogenetic cardiomyopathies Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-350565 SN - 1873-5061 VL - 74 ER - 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 - THES A1 - Horn, Daniela T1 - Kardiotoxizität von CTRPs und das Vorkommen der CTRP-Rezeptoren in Kardiomyozyten T1 - Cardiotoxicity of CTRPs and the presence of CTRP receptors in cardiomyocytes N2 - Die C1q/tumor necrosis factor-related proteins (CTRPs) sind eine Ligandenfamilie aus sezernierten Plasmaproteinen, welche sich in ihrem Grundbauplan ähneln. Daten aus der Literatur deuten darauf hin, dass sie zum Teil positive Effekte auf den Stoffwechsel und das Herz-Kreislaufsystem besitzen und somit eine mögliche therapeutische Zielstruktur darstellen. Während für manche CTRPs bereits Rezeptoren identifiziert werden konnten, ist für andere immer noch nicht geklärt, an welche Rezeptoren sie binden oder über welche sie diese Wirkungen erzielen. Um die CTRPs zukünftig therapeutisch nutzen zu können, muss die Wirkung der CTRPs auf verschiedene Zellen weiter analysiert werden. Dafür wurden in dieser Arbeit Zellen, auf die Expression bereits bekannter CTRP-Rezeptoren hin, untersucht. Des Weiteren wurden die durch CTRP2, CTRP3, CTRP4, CTRP9A, CTRP10, CTRP11, CTRP13 und CTRP14 induzierten Änderungen in der ATP- und Laktatproduktion als Surrogatparameter für Kardiotoxizität in den Kardiomyozytenzelllinien H9c2 und AC16 getestet, um potenziell kardiotoxische Wirkungen frühzeitig erkennen zu können. Es konnte gezeigt werden, dass die CTRPs sicher für Kardiomyozyten zu sein scheinen, was eine wichtige Grundlage für die therapeutische Nutzbarkeit darstellt. N2 - C1q/tumor necrosis factor-related proteins (CTRPs) are a ligand family of secreted plasma proteins that are similar in their basic structure. Literature on the subject indicate that some of them have positive effects on the metabolism and the cardiovascular system and therefore represent a potential therapeutic target structure. While some receptors have already been identified for some CTRPs, for others it is still not clear which receptors they bind to or through which they achieve these effects. In order to be able to use the CTRPs therapeutically in the future, the effect of the CTRPs on different cells must be further analyzed. For that cells were examined in this study for the expression of already known CTRP receptors. Furthermore, CTRP2, CTRP3, CTRP4, CTRP9A, CTRP10, CTRP11, CTRP13 and CTRP14 were tested in the cardiomyocyte cell lines H9c2 and AC16 with respect to their effect on production of ATP and lactate as surrogate parameters for cardiotoxicity in order to be able to recognize potentially cardiotoxic effects at an early stage. It was shown that the CTRPs appear to be safe for cardiomyocytes, which is an important basis for therapeutic utility. KW - Herzmuskelzelle KW - Zelllinie KW - CTRP KW - C1q/tumor necrosis factor-related proteins KW - Kardiomyozyten Y1 - 2024 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-349029 ER -