OXA1L deficiency causes mitochondrial myopathy via reactive oxygen species regulated nuclear factor kappa B signalling pathway

Abstract

Background: OXA1L is crucial for mitochondrial protein insertion and assembly into the inner mitochondrial membrane, and its variants have been recently linked to mitochondrial encephalopathy. However, the definitive pathogenic link between OXA1L variants and mitochondrial diseases as well as the underlying pathogenesis remains elusive.

Methods: In this study, we identified bi-allelic variants of c.620G>T, p.(Cys207Phe) and c.1163_1164del, p.(Val388Alafs*15) in OXA1L gene in a mitochondrial myopathy patient using whole exome sequencing. To unravel the genotype–phenotype relationship and underlying pathogenic mechanism between OXA1L variants and mitochondrial diseases, patient-specific human-induced pluripotent stem cells (hiPSC) were reprogrammed and differentiated into myotubes, while OXA1L knockout human immortalised skeletal muscle cells (IHSMC) and a conditional skeletal muscle knockout mouse model was generated using clustered regularly interspaced short palindromic repeats/Cas9 genomic editing technology.

Results: Both patient-specific hiPSC differentiated myotubes and OXA1L knockout IHSMC showed combined mitochondrial respiratory chain defects and oxidative phosphorylation (OXPHOS) impairments. Notably, in OXA1L-knockout IHSMC, transfection of wild-type human OXA1L but not truncated mutant form rescued the respiratory chain defects. Moreover, skeletal muscle conditional Oxa1l knockout mice exhibited OXPHOS deficiencies and skeletal muscle morphofunctional abnormalities, recapitulating the phenotypes of mitochondrial myopathy. Further functional investigations revealed that impaired OXPHOS resulting of OXA1L deficiency led to elevated reactive oxygen species production, which possibly activated the nuclear factor kappa B signalling pathway, triggering cell apoptosis.

Conclusions: Together, our findings reinforce the genotype–phenotype association between OXA1L variations and mitochondrial diseases and further delineate the potential molecular mechanisms of how OXA1L deficiency causes skeletal muscle deficits in mitochondrial myopathy.

Keywords

mitochondrial myopathy / NF-κB signalling pathway / OXA1L / oxidative phosphorylation / reactive oxygen species

References

[1] Vercellino I, Sazanov LA. The assembly, regulation and function of the mitochondrial respiratory chain. Nat Rev Mol Cell Biol. 2022; 23(2): 141-161.

[2] Bock FJ, Tait SWG. Mitochondria as multifaceted regulators of cell death. Nat Rev Mol Cell Biol. 2020; 21(2): 85-100.

[3] Kaur S, Sharma N, Kumar V, et al. The role of mitophagy in various neurological diseases as a therapeutic approach. Cell Mol Neurobiol. 2023; 43(5): 1849-1865.

[4] Molnar MJ, Kovacs GG. Mitochondrial diseases. Handb Clin Neurol. 2017; 145: 147-155.

[5] Russell OM, Gorman GS, Lightowlers RN, Turnbull DM. Mitochondrial diseases: hope for the future. Cell. 2020; 181(1): 168-188.

[6] Ahmed ST, Craven L, Russell OM, Turnbull DM, Vincent AE. Diagnosis and treatment of mitochondrial myopathies. Neurotherapeutics. 2018; 15(4): 943-953.

[7] Gorman GS, Chinnery PF, DiMauro S, et al. Mitochondrial diseases. Nat Rev Dis Primers. 2016; 2: 16080.

[8] Vafai SB, Mootha VK. Mitochondrial disorders as windows into an ancient organelle. Nature. 2012; 491(7424): 374-383.

[9] Ng YS, Bindoff LA, Gorman GS, et al. Mitochondrial disease in adults: recent advances and future promise. Lancet Neurol. 2021; 20(7): 573-584.

[10] Thompson K, Collier JJ, Glasgow RIC, et al. Recent advances in understanding the molecular genetic basis of mitochondrial disease. J Inherit Metab Dis. 2020; 43(1): 36-50.

[11] Schlieben LD, Prokisch H. Genetics of mitochondrial diseases: current approaches for the molecular diagnosis. Handb Clin Neurol. 2023; 194: 141-165.

[12] Rahman J, Rahman S. Mitochondrial medicine in the omics era. Lancet (London, England). 2018; 391(10139): 2560-2574.

[13] Hennon SW, Soman R, Zhu L, Dalbey RE. YidC/Alb3/Oxa1 family of insertases. J Biol Chem. 2015; 290(24): 14866-14874.

[14] Hildenbeutel M, Theis M, Geier M, et al. The membrane insertase Oxa1 is required for efficient import of carrier proteins into mitochondria. J Mol Biol. 2012; 423(4): 590-599.

[15] Itoh Y, Andréll J, Choi A, et al. Mechanism of membrane-tethered mitochondrial protein synthesis. Science. 2021; 371(6531): 846-849.

[16] Jia L, Dienhart M, Schramp M, McCauley M, Hell K, Stuart RA. Yeast Oxa1 interacts with mitochondrial ribosomes: the importance of the C-terminal region of Oxa1. EMBO J. 2003; 22(24): 6438-6447.

[17] Hell K, Herrmann JM, Pratje E, Neupert W, Stuart RA. Oxa1p, an essential component of the N-tail protein export machinery in mitochondria. Proc Nat Acad Sci USA. 1998; 95(5): 2250-2255.

[18] Hell K, Neupert W, Stuart RA. Oxa1p acts as a general membrane insertion machinery for proteins encoded by mitochondrial DNA. EMBO J. 2001; 20(6): 1281-1288.

[19] Szyrach G, Ott M, Bonnefoy N, Neupert W, Herrmann JM. Ribosome binding to the Oxa1 complex facilitates co-translational protein insertion in mitochondria. EMBO J. 2003; 22(24): 6448-6457.

[20] Stiburek L, Fornuskova D, Wenchich L, Pejznochova M, Hansikova H, Zeman J. Knockdown of human Oxa1l impairs the biogenesis of F1Fo-ATP synthase and NADH:ubiquinone oxidoreductase. J Mol Biol. 2007; 374(2): 506-516.

[21] Thompson K, Mai N, Oláhová M, et al. OXA1L mutations cause mitochondrial encephalopathy and a combined oxidative phosphorylation defect. EMBO Mol Med. 2018; 10(11): e9060.

[22] Frazier AE, Thorburn DR. Biochemical analyses of the electron transport chain complexes by spectrophotometry. Methods Mol Biol. 2012; 837: 49-62.

[23] McCarthy JJ, Srikuea R, Kirby TJ, Peterson CA, Esser KA. Inducible Cre transgenic mouse strain for skeletal muscle-specific gene targeting. Skelet Muscle. 2012; 2(1): 8.

[24] Gu Z, Hübschmann D. simplifyEnrichment: a bioconductor package for clustering and visualizing functional enrichment results. Genom Proteom Bioinf. 2023; 21(1): 190-202.

[25] Morava E, van den Heuvel L, Hol F, et al. Mitochondrial disease criteria: diagnostic applications in children. Neurology. 2006; 67(10): 1823-1826.

[26] Parikh S, Goldstein A, Koenig MK, et al. Diagnosis and management of mitochondrial disease: a consensus statement from the Mitochondrial Medicine Society. Genet Med. 2015; 17(9): 689-701.

[27] Romero-Moya D, Santos-Ocaña C, Castaño J, et al. Genetic rescue of mitochondrial and skeletal muscle impairment in an induced pluripotent stem cells model of coenzyme Q(10) deficiency. Stem Cells. 2017; 35(7): 1687-1703.

[28] Segalés J, Perdiguero E, Muñoz-Cánoves P. Epigenetic control of adult skeletal muscle stem cell functions. FEBS J. 2015; 282(9): 1571-1588.

[29] Shadel GS, Horvath TL. Mitochondrial ROS signaling in organismal homeostasis. Cell. 2015; 163(3): 560-569.

[30] Dong W, Chen W, Zou H, et al. Ginsenoside Rb1 prevents oxidative stress-induced apoptosis and mitochondrial dysfunction in muscle stem cells via NF-κB pathway. Oxid Med Cell Long. 2022; 2022: 9159101.

[31] Morgan MJ, Liu ZG. Crosstalk of reactive oxygen species and NF-κB signaling. Cell Res. 2011; 21(1): 103-115.

[32] Dogan SA, Giacchin G, Zito E, Viscomi C. Redox signaling and stress in inherited myopathies. Antioxid Redox Signal. 2022; 37(4-6): 301-323.

[33] Ronchi D, Garbellini M, Magri F, et al. A biallelic variant in COX18 cause isolated Complex IV deficiency associated with neonatal encephalo-cardio-myopathy and axonal sensory neuropathy. Eur J Hum Genet. 2023; 31(12): 1414-1420.

[34] Mavillard F, Guerra-Castellano A, Guerrero-Gómez D, et al. A splice-altering homozygous variant in COX18 causes severe sensory-motor neuropathy with oculofacial apraxia. Biochim Biophys Acta Mol Basis Dis. 2024; 1870(7): 167330.

[35] Armirola-Ricaurte C, Morant L, Adant I, et al. Biallelic variants in COX18 cause a mitochondrial disorder primarily manifesting as peripheral neuropathy. medRxiv. 2024.

[36] Vögtle FN, Brändl B, Larson A, et al. Mutations in PMPCB encoding the catalytic subunit of the mitochondrial presequence protease cause neurodegeneration in early childhood. Am J Hum Genet. 2018; 102(4): 557-573.

[37] Yoon JY, Daneshgar N, Chu Y, et al. Metabolic rescue ameliorates mitochondrial encephalo-cardiomyopathy in murine and human iPSC models of Leigh syndrome. Clin Transl Med. 2022; 12(7): e954.

[38] Bitto A, Grillo AS, Ito TK, et al. Acarbose suppresses symptoms of mitochondrial disease in a mouse model of Leigh syndrome. Nat Metab. 2023; 5(6): 955-967.

[39] Ruzzenente B, Rötig A, Metodiev MD. Mouse models for mitochondrial diseases. Hum Mol Genet. 2016; 25(R2): R115-R122.

[40] van de Wal MAE, Adjobo-Hermans MJW, Keijer J, et al. Ndufs4 knockout mouse models of Leigh syndrome: pathophysiology and intervention. Brain: J Neurol. 2022; 145(1): 45-63.

[41] Di Leo V, Bernardino Gomes TM, Vincent AE. Interactions of mitochondrial and skeletal muscle biology in mitochondrial myopathy. Biochem J. 2023; 480(21): 1767-1789.

[42] Auré K, Fayet G, Leroy JP, Lacène E, Romero NB, Lombès A. Apoptosis in mitochondrial myopathies is linked to mitochondrial proliferation. Brain: J Neurol. 2006; 129(Pt 5): 1249-1259.

[43] Barbieri E, Sestili P. Reactive oxygen species in skeletal muscle signaling. J Signal Transduct. 2012; 2012: 982794.

[44] Gallego-Selles A, Galvan-Alvarez V, Martinez-Canton M, et al. Fast regulation of the NF-κB signalling pathway in human skeletal muscle revealed by high-intensity exercise and ischaemia at exhaustion: role of oxygenation and metabolite accumulation. Redox Biol. 2022; 55: 102398.

[45] Zhou T, Prather ER, Garrison DE, Zuo L. Interplay between ROS and antioxidants during ischemia-reperfusion injuries in cardiac and skeletal muscle. Int J Mol Sci. 2018; 19(2): 417.

[46] Haramizu S, Asano S, Butler DC, et al. Dietary resveratrol confers apoptotic resistance to oxidative stress in myoblasts. J Nutr Biochem. 2017; 50: 103-115.

[47] Lee S, Shin HS, Shireman PK, Vasilaki A, Van Remmen H, Csete ME. Glutathione-peroxidase-1 null muscle progenitor cells are globally defective. Free Radical Biol Med. 2006; 41(7): 1174-1184.

[48] Choi MH, Ow JR, Yang ND, Taneja R. Oxidative stress-mediated skeletal muscle degeneration: molecules, mechanisms, and therapies. Oxid Med Cell Long. 2016; 2016: 6842568.

[49] Ardite E, Barbera JA, Roca J, Fernández-Checa JC. Glutathione depletion impairs myogenic differentiation of murine skeletal muscle C2C12 cells through sustained NF-kappaB activation. Am J Pathol. 2004; 165(3): 719-728.

[50] Catani MV, Savini I, Duranti G, et al. Nuclear factor kappaB and activating protein 1 are involved in differentiation-related resistance to oxidative stress in skeletal muscle cells. Free Radical Biol Med. 2004; 37(7): 1024-1036.

[51] Guttridge DC, Mayo MW, Madrid LV, Wang CY. NF-kappaB-induced loss of MyoD messenger RNA: possible role in muscle decay and cachexia. Science. 2000; 289(5488): 2363-2366.

[52] Wang H, Hertlein E, Bakkar N, et al. NF-kappaB regulation of YY1 inhibits skeletal myogenesis through transcriptional silencing of myofibrillar genes. Mol Cell Biol. 2007; 27(12): 4374-4387.

[53] Baechler BL, Bloemberg D, Quadrilatero J. Mitophagy regulates mitochondrial network signaling, oxidative stress, and apoptosis during myoblast differentiation. Autophagy. 2019; 15(9): 1606-1619.

[54] Chabi B, Hennani H, Cortade F, Wrutniak-Cabello C. Characterization of mitochondrial respiratory complexes involved in the regulation of myoblast differentiation. Cell Biol Int. 2021; 45(8): 1676-1684.

[55] Wagatsuma A, Sakuma K. Mitochondria as a potential regulator of myogenesis. Sci World J. 2013; 2013: 593267.

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2025 The Author(s). Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.