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Mitochondrial “dysmorphology” in variant classification

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Mitochondrial disorders are challenging to diagnose. Exome sequencing has greatly enhanced the diagnostic precision of these disorders although interpreting variants of uncertain significance (VUS) remains a formidable obstacle. Whether specific mitochondrial morphological changes can aid in the classification of these variants is unknown. Here, we describe two families (four patients), each with a VUS in a gene known to affect the morphology of mitochondria through a specific role in the fission–fusion balance. In the first, the missense variant in MFF, encoding a fission factor, was associated with impaired fission giving rise to a characteristically over-tubular appearance of mitochondria. In the second, the missense variant in DNAJA3, which has no listed OMIM phenotype, was associated with fragmented appearance of mitochondria consistent with its published deficiency states. In both instances, the highly specific phenotypes allowed us to upgrade the classification of the variants. Our results suggest that, in select cases, mitochondrial “dysmorphology” can be helpful in interpreting variants to reach a molecular diagnosis.

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  • Alkuraya FS (2010) Autozygome decoded. Genet Med 12:765–771

    Article  Google Scholar 

  • Alkuraya FS (2013) The application of next-generation sequencing in the autozygosity mapping of human recessive diseases. Hum Genet 132:1197–1211

    Article  CAS  Google Scholar 

  • Bartsakoulia M, Pyle A, Troncoso-Chandia D, Vial-Brizzi J, Paz-Fiblas MV, Duff J, Griffin H, Boczonadi V, Lochmuller H, Kleinle S, Chinnery PF, Grunert S, Kirschner J, Eisner V, Horvath R (2018) A novel mechanism causing imbalance of mitochondrial fusion and fission in human myopathies. Hum Mol Genet 27:1186–1195.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bhattacharyya T, Karnezis AN, Murphy SP, Hoang T, Freeman BC, Phillips B, Morimoto RI (1995) Cloning and subcellular localization of human mitochondrial hsp70. J Biol Chem 270:1705–1710

    Article  CAS  Google Scholar 

  • Campello S, Lacalle RA, Bettella M, Mañes S, Scorrano L, Viola A (2006) Orchestration of lymphocyte chemotaxis by mitochondrial dynamics. J Exp Med 203:2879–2886

    Article  CAS  Google Scholar 

  • Chen H, Vermulst M, Wang YE, Chomyn A, Prolla TA, McCaffery JM, Chan DC (2010) Mitochondrial fusion is required for mtDNA stability in skeletal muscle and tolerance of mtDNA mutations. Cell 141:280–289

    Article  CAS  Google Scholar 

  • Collins TJ, Berridge MJ, Lipp P, Bootman MD (2002) Mitochondria are morphologically and functionally heterogeneous within cells. EMBO J 21:1616–1627

    Article  CAS  Google Scholar 

  • Elwi AN, Lee B, Meijndert HC, Braun JE, Kim S-W (2012) Mitochondrial chaperone DnaJA3 induces Drp1-dependent mitochondrial fragmentation. Int J Biochem Cell Biol 44:1366–1376

    Article  CAS  Google Scholar 

  • Feoktistova M, Geserick P, Leverkus M (2016) Crystal violet assay for determining viability of cultured cells. Cold Spring Harb Protoc 2016:pdb.prot087379

    Article  Google Scholar 

  • Frank S, Gaume B, Bergmann-Leitner ES, Leitner WW, Robert EG, Catez F, Smith CL, Youle RJ (2001) The role of dynamin-related protein 1, a mediator of mitochondrial fission, in apoptosis. Dev Cell 1:515–525

    Article  CAS  Google Scholar 

  • Frezza C, Cipolat S, De Brito OM, Micaroni M, Beznoussenko GV, Rudka T, Bartoli D, Polishuck RS, Danial NN, De Strooper B (2006) OPA1 controls apoptotic cristae remodeling independently from mitochondrial fusion. Cell 126:177–189

    Article  CAS  Google Scholar 

  • Gai X, Ghezzi D, Johnson MA, Biagosch CA, Shamseldin HE, Haack TB, Reyes A, Tsukikawa M, Sheldon CA, Srinivasan S (2013) Mutations in FBXL4, encoding a mitochondrial protein, cause early-onset mitochondrial encephalomyopathy. Am J Hum Genet 93:482–495

    Article  CAS  Google Scholar 

  • Goswami AV, Chittoor B, D’Silva P (2010) Understanding the functional interplay between mammalian mitochondrial Hsp70 chaperone machine components. J Biol Chem 285:19472–19482

    Article  CAS  Google Scholar 

  • Green ED, Gunter C, Biesecker LG, Di Francesco V, Easter CL, Feingold EA, Felsenfeld AL, Kaufman DJ, Ostrander EA, Pavan WJ (2020) Strategic vision for improving human health at The Forefront of Genomics. Nature 586:683–692

    Article  CAS  Google Scholar 

  • Hayashi M, Imanaka-Yoshida K, Yoshida T, Wood M, Fearns C, Tatake RJ, Lee J-D (2006) A crucial role of mitochondrial Hsp40 in preventing dilated cardiomyopathy. Nat Med 12:128–132

    Article  CAS  Google Scholar 

  • Hsieh T-C, Mensah MA, Pantel JT, Aguilar D, Bar O, Bayat A, Becerra-Solano L, Bentzen HB, Biskup S, Borisov O (2019) PEDIA: prioritization of exome data by image analysis. Genet Med 21:2807–2814

    Article  Google Scholar 

  • Iosefson O, Azem A (2012) Many faces of Mortalin and Tid1. In: Kaul S, Wadhwa R (eds) Mortalin biology: life, stress and death. Springer, Dordrecht.

  • Kampinga HH, Craig EA (2010) The HSP70 chaperone machinery: J proteins as drivers of functional specificity. Nat Rev Mol Cell Biol 11:579–592

    Article  CAS  Google Scholar 

  • Koch J, Feichtinger RG, Freisinger P, Pies M, Schrödl F, Iuso A, Sperl W, Mayr JA, Prokisch H, Haack TB (2016) Disturbed mitochondrial and peroxisomal dynamics due to loss of MFF causes Leigh-like encephalopathy, optic atrophy and peripheral neuropathy. J Med Genet 53:270–278

    Article  CAS  Google Scholar 

  • Linnoila J, Wang Y, Yao Y, Wang Z-Z (2008) A mammalian homolog of Drosophila tumorous imaginal discs, Tid1, mediates agrin signaling at the neuromuscular junction. Neuron 60:625–641

    Article  CAS  Google Scholar 

  • Lo J-F, Hayashi M, Woo-Kim S, Tian B, Huang J-F, Fearns C, Takayama S, Zapata JM, Yang Y, Lee J-D (2004) Tid1, a cochaperone of the heat shock 70 protein and the mammalian counterpart of the Drosophila tumor suppressor l (2) tid, is critical for early embryonic development and cell survival. Mol Cell Biol 24:2226–2236

    Article  CAS  Google Scholar 

  • Mai S, Klinkenberg M, Auburger G, Bereiter-Hahn J, Jendrach M (2010) Decreased expression of Drp1 and Fis1 mediates mitochondrial elongation in senescent cells and enhances resistance to oxidative stress through PINK1. J Cell Sci 123:917–926

    Article  CAS  Google Scholar 

  • Mistry J, Chuguransky S, Williams L, Qureshi M, Salazar GA, Sonnhammer ELL, Tosatto SCE, Paladin L, Raj S, Richardson LJ, Finn RD, Bateman A (2021) Pfam: the protein families database in 2021. Nucleic Acids Res 49:D412–D419.

    Article  CAS  PubMed  Google Scholar 

  • Monies D, Abouelhoda M, AlSayed M, Alhassnan Z, Alotaibi M, Kayyali H, Al-Owain M, Shah A, Rahbeeni Z, Al-Muhaizea MA (2017) The landscape of genetic diseases in Saudi Arabia based on the first 1000 diagnostic panels and exomes. Hum Genet 136:921–939

    Article  CAS  Google Scholar 

  • Nasca A, Nardecchia F, Commone A, Semeraro M, Legati A, Garavaglia B, Ghezzi D, Leuzzi V (2018) Clinical and biochemical features in a patient with mitochondrial fission factor gene alteration. Front Genet 9:625.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ng AC-H, Baird SD, Screaton RA (2014) Essential role of TID1 in maintaining mitochondrial membrane potential homogeneity and mitochondrial DNA integrity. Mol Cell Biol 34:1427–1437

    Article  Google Scholar 

  • Panda I, Ahmad I, Sagar S, Zahra S, Shamim U, Sharma S, Faruq M (2020) Encephalopathy due to defective mitochondrial and peroxisomal fission 2 caused by a novel MFF gene mutation in a young child. Clin Genet 97:933–937

    Article  CAS  Google Scholar 

  • Patra M, Weiss C, Abu-Libdeh B, Ashhab M, Abuzer S, Elpeleg O, Mahajnah M, Kessel A, Azem A (2019) A novel variant of the human mitochondrial DnaJ protein, Tid1, associates with a human disease exhibiting developmental delay and polyneuropathy. Eur J Hum Genet 27:1072–1080

    Article  Google Scholar 

  • Richards S, Aziz N, Bale S, Bick D, Das S, Gastier-Foster J, Grody WW, Hegde M, Lyon E, Spector E (2015) Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology. Genet Med 17:405–423

    Article  Google Scholar 

  • Scorrano L, Ashiya M, Buttle K, Weiler S, Oakes SA, Mannella CA, Korsmeyer SJ (2002) A distinct pathway remodels mitochondrial cristae and mobilizes cytochrome c during apoptosis. Dev Cell 2:55–67

    Article  CAS  Google Scholar 

  • Shamseldin HE, Alshammari M, Al-Sheddi T, Salih MA, Alkhalidi H, Kentab A, Repetto GM, Hashem M, Alkuraya FS (2012) Genomic analysis of mitochondrial diseases in a consanguineous population reveals novel candidate disease genes. J Med Genet 49:234–241

    Article  Google Scholar 

  • Shamseldin HE, Smith LL, Kentab A, Alkhalidi H, Summers B, Alsedairy H, Xiong Y, Gupta VA, Alkuraya FS (2016) Mutation of the mitochondrial carrier SLC25A42 causes a novel form of mitochondrial myopathy in humans. Hum Genet 135:21–30

    Article  CAS  Google Scholar 

  • Shamseldin HE, Alasmari A, Salih MA, Samman MM, Mian SA, Alshidi T, Ibrahim N, Hashem M, Faqeih E, Al-Mohanna F (2017) A null mutation in MICU2 causes abnormal mitochondrial calcium homeostasis and a severe neurodevelopmental disorder. Brain 140:2806–2813

    Article  Google Scholar 

  • Shen JJ, Wortmann SB, de Boer L, Kluijtmans LA, Huigen MC, Koch J, Ross S, Collins CD, van der Lee R, van Karnebeek CD (2020) The role of clinical response to treatment in determining pathogenicity of genomic variants. Genet Med 23(3):581-585

  • Syken J, De-Medina T, Münger K (1999) TID1, a human homolog of the Drosophila tumor suppressor l (2) tid, encodes two mitochondrial modulators of apoptosis with opposing functions. Proc Natl Acad Sci 96:8499–8504

    Article  CAS  Google Scholar 

  • Thorburn D (2004) Mitochondrial disorders: prevalence, myths and advances. J Inherit Metab Dis 27:349–362

    Article  CAS  Google Scholar 

  • Valente AJ, Maddalena LA, Robb EL, Moradi F, Stuart JA (2017) A simple ImageJ macro tool for analyzing mitochondrial network morphology in mammalian cell culture. Acta Histochem 119:315–326

    Article  CAS  Google Scholar 

  • Zhao Q, Wang J, Levichkin IV, Stasinopoulos S, Ryan MT, Hoogenraad NJ (2002) A mitochondrial specific stress response in mammalian cells. EMBO J 21:4411–4419

    Article  CAS  Google Scholar 

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We would like to thank the study families for their enthusiastic participation. We acknowledge the technical help by the Sequencing, Genotyping and Bioinformatics Core Facilities at KFSHRC.

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Correspondence to Fowzan S. Alkuraya.

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Supplementary file1 Fig. S1 A and B Cell viability of patients (family 1-IV:2 and family 2-IV:3, respectively), is significantly affected compared to controls (experimental and biological triplicates, and unpaired t-test used for statistically analysis). Fig. S2 Mitochondria network analysis tool (MiNA) applied for analysis of MFF-defective cells (family 2-IV:3) compared to controls shows long tangled patient mitochondria, with statistically significant difference in footprint. Fig. S3 Immunoblot results for A MFF protein in (family 2-IV:3) and B DNAJA3 protein in (family 1-IV:2) show no significant difference in the protein level extracted from the affected fibroblasts compared to the control (representative of biological triplicates). (PPTX 937 KB)

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Shamseldin, H.E., Alhashem, A., Tabarki, B. et al. Mitochondrial “dysmorphology” in variant classification. Hum Genet 141, 55–64 (2022).

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