Mitochondrial defect in muscle precedes neuromuscular junction degeneration and motor neuron death in CHCHD10S59L/+ mouse
Recently, we provided genetic basis showing that mitochondrial dysfunction can trigger motor neuron degeneration, through identification of CHCHD10 encoding a mitochondrial protein. We reported patients, carrying the p.Ser59Leu heterozygous mutation in CHCHD10, from a large family with a mitochondrial myopathy associated with motor neuron disease (MND). Rapidly, our group and others reported CHCHD10 mutations in amyotrophic lateral sclerosis (ALS), frontotemporal dementia-ALS and other neurodegenerative diseases. Here, we generated knock-in (KI) mice, carrying the p.Ser59Leu mutation, that mimic the mitochondrial myopathy with mtDNA instability displayed by the patients from our original family. Before 14 months of age, all KI mice developed a fatal mitochondrial cardiomyopathy associated with enhanced mitophagy. CHCHD10S59L/+ mice also displayed neuromuscular junction (NMJ) and motor neuron degeneration with hyper-fragmentation of the motor end plate and moderate but significant motor neuron loss in lumbar spinal cord at the end stage of the disease. At this stage, we observed TDP-43 cytoplasmic aggregates in spinal neurons. We also showed that motor neurons differentiated from human iPSC carrying the p.Ser59Leu mutation were much more sensitive to Staurosporine or glutamate-induced caspase activation than control cells. These data confirm that mitochondrial deficiency associated with CHCHD10 mutations can be at the origin of MND. CHCHD10 is highly expressed in the NMJ post-synaptic part. Importantly, the fragmentation of the motor end plate was associated with abnormal CHCHD10 expression that was also observed closed to NMJs which were morphologically normal. Furthermore, we found OXPHOS deficiency in muscle of CHCHD10S59L/+ mice at 3 months of age in the absence of neuron loss in spinal cord. Our data show that the pathological effects of the p.Ser59Leu mutation target muscle prior to NMJ and motor neurons. They likely lead to OXPHOS deficiency, loss of cristae junctions and destabilization of internal membrane structure within mitochondria at motor end plate of NMJ, impairing neurotransmission. These data are in favor with a key role for muscle in MND associated with CHCHD10 mutations.
KeywordsCHCHD10 Mitochondrial disorder ALS Mouse model iPSC
We thank our colleagues from Genetic Engineering and Mouse Transgenesis department at service unit CIPHE (Centre d’Immunophénomique, Aix Marseille Université, INSERM, CNRS, Phenomin, Marseille, France) and mainly Frederic Fiore who provided expertise and know-how and perform the generation of the CHCHD10S59L/+ mice that greatly assisted our project. We also thank our colleagues, including Benoit Petit-Demoulière, Ghina Bou-About, Hamid Meziane, Hugues Jacobs, Marie-France Champy, Patrick Reilly, Tania Sorg and Yann Herault from Phenomin (Institut Clinique de la souris, 1 rue Laurent Fries, 67404 ILLKIRCH cedex 2—CNRS, UMR7104, Illkirch, France—INSERM, U964, Illkirch, France—Université de Strasbourg, France) for their help in phenotyping mice. We gratefully acknowledge the IRCAN’s Molecular and Cellular Core Imaging (PICMI) facility, supported financially by FEDER, Conseil régional Provence Alpes-Côte d’Azur, Conseil Départemental 06, Cancéropôle PACA, Gis Ibisa and Inserm, the IRCAN’s Animal core facility, supported by Région Provence Alpes-Côte d’Azur and Inserm, the IRCAN’s Histology core facility, supported by Région Provence Alpes-Côte d’Azur and Université de Nice Sophia-Antipolis, the Centre Méditerranéen de Médecine Moléculaire (C3 M) imaging facility, the University’s CCMA Electron Microscopy facility supported by Université de Nice Sophia-Antipolis, Région Provence Alpes-Côte d’Azur, Conseil Départemental 06, and Gis Ibisa and, the ICM CELIS-iPS internal platform. We also thank Dr Luisa Villa and Dr Pamela Moceri for their help in interpreting muscle and cardiac results, respectively, and Sandra Foustoul and Alyssia Mari for technical help.
ECG and SBa designed, performed experiments and analyzed data on mouse models, and contributed to the main text. BMH designed, performed experiments and analyzed data on cell models (NSC34 and iPSCs-derived motor neurons), and contributed to the main text. KF designed, performed experiments and analyzed data on respiratory chain experiments. SL-G performed electron microscopy experiments. JN, AM-C, FL, BR, GA and CC performed and analyzed experiments on mouse models. SBi generated iPSC. CL-O and DB provided the motor neuron differentiation protocol and advices on the iPSC daily culture and motor neuron differentiation. DB provides advices and helped in data interpretation on iPSC-derived motor neuron experiments and contributed to the main text. AC provided technical advices for motor neurons staining and count. FM helped with the interpretation of metabolic data. SBo and CL provided advices for the NMJ and motor neurons analyses and helped in data interpretation. J‐ER designed and analyzed data on cell models and contributed to the main text. VP‐F designed, supervised the project and drafted the manuscript. All the authors read and approved the final submission.
This work was made possible by Grants to VP-F from the ANR (Agence Nationale de la Recherche) ANR-16-CE16-0024-01, from the AFM-Téléthon (Association Française contre les Myopathies) #20947 and from the Fondation Maladies Rares. The research leading to the iPSC results has received funding from the program “Investissements d’avenir” ANR-11-INBS-0011-NeurATRIS: Translational Research Infrastructure for Biotherapies in Neurosciences”. ECG and BMH are postdoctoral fellows supported by ANR-16-CE16-0024-01. CL-O is a Ph.D. fellow supported by ANR-10-LABX-73.
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Conflict of interest
No conflict of interest.
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