Anand, G

Barbosa, IA

Carver, J

Dombi, E

Fratter, C

Harrison, V

Hofer, M

Jayawant, S

Ladds, E

Poulton, J

Simpson, MA

Whitney, A

Hofer, Monika

Anand, Getha

Carver, Janet

Jayawant, Sandeep

Poulton, Joanna

Oxford University Research Archive

CLINICAL/SCIENTIFIC NOTES OPEN ACCESSDe novo DNM1L mutation associated withmitochondrial epilepsy syndrome with feversensitivityEmma Ladds, MBBCh, Andrea Whitney, MB, BS, Eszter Dombi, BSc, Monika Hofer, BMSc, MBChB,Geetha Anand, MRCPCH, MD, Victoria Harrison, MBChB, Carl Fratter, MPhil, MChem, Janet Carver, MSc,Ines A. Barbosa, DPhil,Michael Simpson, PhD, Sandeep Jayawant,MD, FRCPCH, and JoannaPoulton, DM, FRCPNeurol Genet 2018;4:e258. doi:10.1212/NXG.0000000000000258CorrespondenceDr. PoultonJoanna.poulton@wrh.ox.ac.ukCatastrophic epileptic encephalopathy of unclear etiology following a mild metabolic insultgenerally has a poor outcome. Here, we present 2 such unrelated individuals in whom whole-exome sequencing identified the same de novo recurrent mutation (c.1207C>T p.Arg403Cys)in the gene encoding the guanosine triphosphatase (GTPase) Dynamin-1 like Protein(DNM1L) (reference sequence NM_012062.4).The dynamic fission and fusion of the intracellular mitochondrial network are essential tofacilitatemitophagy and thusmitochondrial quality and function.1 Duringmitochondrial division,the GTPaseDNM1L forms multimeric collars at specific fission sites, constricting portions of themitochondrial reticulum and generating fragments for engulfment and degradation.2DNM1L has been implicated in several presentations of refractory epilepsy.3 Both of ourpatients exhibited signs of preexisting developmental delay and presented with epilepsy during,or recently following, a febrile illness or exercise. Elevated lactate levels, epilepsia partialiscontinua, nonspecific imaging, and evidence of lipid storage myopathy all support mitochon-drial dysfunction (See table for presentation summary and e-case report for details, links.lww.com/NXG/A63). This evidence supports an etiological role for DNM1L in mitochondrialepilepsy syndrome with fever sensitivity (MEFS).MethodsWe used IN Cell Analyzer 1000 (IN Cell 1000), a previously validated high-throughput imagingmethod for quantifying mitophagy and mitochondrial DNA (mtDNA) in cultured fibroblastsfrom patients compared with cultures derived from karyotypically normal controls. Cells wereimmunostained for the autophagy marker Light Chain 3 (LC3) and the mitochondrial importreceptor, translocase of outer membrane 20 (TOM20) and analyzed with IN Cell1000. Thereadout for mitophagy was colocalization of LC3 puncta with TOM20-positive mitochondria.ResultsWe showed that the mitochondria in fibroblasts from both patients are lengthened andhyperpolarized (figure e-1, A and B, links.lww.com/NXG/A62). Total mitophagic flux wasincreased, showing that mitophagy is activated (figure e-1C, i). This is consistent with theFrom the Harvard Chan School of Public Health (E.L.), Harvard University, Boston, MA; Department of Paediatrics (A.W.), University Hospital Southampton NHS Foundation Trust;Nuffield Department Women’s + Reproductive Health (E.D., J.C., J.P.), University of Oxford, The Women’s Centre; Department of Neuropathology (M.H.), Oxford University HospitalsNHS Foundation Trust; Oxford Children’s Hospital (G.A., S.J.), Oxford University Hospitals NHS Foundation Trust; Wessex Clinical Genetics Service (V.H.), University Hospital South-ampton NHS Foundation Trust; and Department of Medical and Molecular Genetics (C.F., I.A.B., M.S.), King’s College London School of Basic and Medical Biosciences, London, UnitedKingdom.Funding information and disclosures are provided at the end of the article. Full disclosure form information provided by the authors is available with the full text of this article atNeurology.org/NG.The Article Processing Charge was funded by the authors.This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND), which permits downloadingand sharing the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal.Copyright © 2018 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology. 1mtDNA depletion documented in fibroblasts (figure e-1C, iii)and the borderline low mtDNA content in skeletal musclefrom patient 1 (table).DiscussionIn 2 patients, we identified a de novo dominant mutation inDMN1L, the same mutation having now been identified in 4unrelated patients with refractory epilepsy.3 Presentationfeatures and investigation findings supported an underlyingmitochondrial pathology. Altered mitochondrial dynamics arenow a well-established cause of disease (see supplementaryinformation for reference, links.lww.com/NXG/A63), anddefective mitochondrial fission may both cause synaptic dys-function1 and impair responses to infection.4In mice, constitutive homozygous knockouts of Drp1 (themurine homolog of DNM1L) do not survive embryogenesis,while conditional ablation leads to developmental defects,both associated with abnormal mitochondrial fission.5 Indi-viduals heterozygous for DNM1L p.Arg403Cys displaya milder phenotype, both in terms of mitochondrial structuraland functional abnormalities and symptom severity.Previously described cases similarly display several years ofrelatively normal development followed by severe, refractoryepilepsy following a mild metabolic insult, vaccinations, or low-grade fever, resulting in profound global developmental delay.3As in case 1, nonspecific thalamic hyperintensities were seen onMRI scans on the 2 previously reported probands with thep.Arg403Cys mutation.3 Case 2 demonstrated diffuse cerebralvolume atrophy. These changes are in keeping with previouslyreported nonspecific T2 MRI hyperintensities and cerebral orcerebellar atrophy seen in other mitochondrial disorders. Itremains to be seen whether MEFS is part of the same clinicalspectrum of other conditions associated with status epilepticusrelated to febrile illnesses, i.e., new-onset refractory status epi-lepticus or febrile illness–related epilepsy syndrome.6DNM1L is required for division of mitochondria and perox-isomes, interacting with receptor Mff and endoplasmic re-ticulum (ER) components. The missense mutation shared inTable Comparison of clinical presentations and investigationsCase 1 Case 2Prepresentation development Moderate language delay Mild general developmental delaySeizure features Age 3: focal status epilepticus; continuous partialseizures and a right hemiparesisAge 5: asymmetric generalized tonic-clonicseizures provoked by febrile illness.Age 6: focal and convulsive status epilepticusPersistent right-sided clonic and occasional non-or generalized convulsive status epilepticusEtiological insult 5 days after influenza vaccination; concurrentmild febrile illnessShortly after febrile illness or exerciseBiochemical findings Increased lactate level (2.7 mmol/L) Increased lactate level (2.5 mmol/L)EEG findings Left frontotemporal eliptogenic focus Focal right-sided clonic seizures associated withleft frontal ictal onset; nonconvulsive statusepilepticus with bilateral slow spike wavesMRI findings Transient left thalamic lesion with progression ofT2 white matter hyperintensityNormal morphology and myelination; globalbilateral cerebral volume lossMuscle biopsy Variable muscle fiber size and type II atrophy Type II atrophy and cytochrome oxidase negative(COX-negative) succinate dehydrogenase-positive (SDH-positive) fibersMuscle lipid Increased Prominent in type I fibersMuscle mtDNA 33% of expected (borderline low) 71% of expected (normal)Mitochondrial appearance Prominent—consistent with long mitochondriain fibroblastsProminent—consistent with long mitochondriain fibroblastsMitochondrial activity Borderline low complex IV NormalWhole mitochondrial genome sequencing No pathologic variants No pathologic variantsGenes implicated in autosomal mitochondrialdisordersNo abnormalities in 17 genes Normal NPC1, NPC2, and POLGWhole-exome sequencing c.1207C>T p.Arg403Cys in DNML1 c.1207C>T p.Arg403Cys in DNML1Treatment Moderate response to carbamazepine,clobazam, and coenzyme QModerate response to levetiracetam,carbamazepine, and clobazam2 Neurology: Genetics | Volume 4, Number 4 | August 2018 Neurology.org/NGour cases lies in the middle domain of DNM1L,3 impairingoligomerization and recruitment to mitochondria3 consistentwith our findings of elongated mitochondria in fibroblastsfrom both patients (figure e-1A, links.lww.com/NXG/A62).Furthermore, knockdown of the DNM1L ortholog Drp1 inmouse cells and its ligand Mff can each cause mtDNA de-pletion and mitochondrial dysfunction,7 consistent with theborderline low mtDNA content and COX activity in skeletalmuscle we demonstrated in case 1. This is likely due to in-creased mitophagic flux (figure e-1C, i).Our findings suggest that DNM1L is implicated as a geneticcontributor to MEFS. DNM1L p.Arg403Cys mutationscreening could therefore be useful in patients with similarpresentations and in identifying impaired mitochondrial fis-sion causing synaptic dysfunction1 and defective response toinfection.Author contributionsE. Ladds wrote the initial case reports and manuscript draft,was part of the editing and submissions process, and preparedthe cases for presentation at the British Paediatric NeurologyAssociation Conference 2018. A. Whitney was the pediatri-cian responsible for identifying SP and summarizing his case.E. Dombi and M. Hofer performed the laboratory tests andanalysis of the data. G. Anand was the pediatrician responsiblefor initially summarizing MN’s clinical presentation with thekey learning points, bringing together the coauthors, andinitiating and supervising case presentation at the NationalBritish Paediatric Neurology Association Conference 2018.He also contributed to the final editing process. V. Harrisoninitiated the laboratory tests and analysis of the data. C.Fratter, J. Carver, and I.A. Barbosa performed the laboratorytests and analysis of the data. M. Simpson performed thewhole-exome sequencing. S. Jayawant was the pediatric neu-rology consultant responsible for MN’s clinical care andidentifying the case for publication. J. Poulton performedconceptualization of the study and analysis of the data andrevised the manuscript.Study fundingThe authors acknowledge Kate Sergeant and Charu Desh-pande for their involvement in whole-exome sequencing andparental testing of case 1, which was supported by the LilyFoundation.DisclosureE. Ladds reports no disclosures. A Whitney has served on thescientific advisory board of Zogenix and has received fundingfor speaker honoraria from Zogenix. E. Dombi, M. Hofer, G.Anand, V. Harrison, C. Fratter, and J. Carver report no dis-closures. I.A. Barbosa has received research funding from theLily Foundation. M. Simpson is employed by Genomics Plc.S. Jayawant reports no disclosures. J. Poulton receives re-search support from the Lily Foundation, the WellcomeTrust, and the UK Medical Research Council. Full disclosureform information provided by the authors is available with thefull text of this article at Neurology.org/NG.Received February 5, 2018. Accepted in final form May 23, 2018.References1. Itoh K, Nakamura K, Iijima M, Sesaki H. Mitochondrial dynamics in neuro-degeneration. Trends Cell Biol 2013;23:64–71.2. Smirnova E, Griparic L, Shurland DL, van der Bliek AM. Dynamin-related proteinDrp1 is required for mitochondrial division in mammalian cells. Mol Biol Cell 2001;12:2245–2256.3. Fahrner JA, Liu R, Perry MS, Klein J, Chan DC. A novel de novo dominant negativemutation in DNM1L impairs mitochondrial fission and presents as childhood epi-leptic encephalopathy. Am J Med Genet A 2016;170:2002–2011.4. Shahni R, Cale CM, Anderson G, et al. Signal transducer and activator of transcription2 deficiency is a novel disorder of mitochondrial fission. Brain 2015;138:2834–2846.5. Cahill TJ, Leo V, Kelly M, et al. Resistance of dynamin-related protein 1 oligomers todisassembly impairs mitophagy, resulting in myocardial inflammation and heart fail-ure. J Biol Chem 2015;290:25907–25919.6. Gaspard N, Hirsch LJ, Sculier C, et al. New-onset refractory status epilepticus(NORSE) and febrile infection-related epilepsy syndrome (FIRES): state of the artand perspectives. Epilepsia 2018;59:745–752.7. Parone PA, Da Cruz S, Tondera D, et al. Preventing mitochondrial fission impairs mito-chondrial function and leads to loss of mitochondrial DNA. PLoS One 2008;3:e3257.Neurology.org/NG Neurology: Genetics | Volume 4, Number 4 | August 2018 3DOI 10.1212/NXG.00000000000002582018;4; Neurol Genet Emma Ladds, Andrea Whitney, Eszter Dombi, et al. sensitivity mutation associated with mitochondrial epilepsy syndrome with feverDNM1LDe novo This information is current as of August 3, 2018ServicesUpdated Information & http://ng.neurology.org/content/4/4/e258.full.htmlincluding high resolution figures, can be found at:References http://ng.neurology.org/content/4/4/e258.full.html##ref-list-1This article cites 7 articles, 2 of which you can access for free at: Subspecialty Collections http://ng.neurology.org//cgi/collection/mitochondrial_disordersMitochondrial disorders http://ng.neurology.org//cgi/collection/genetic_linkageGenetic linkage http://ng.neurology.org//cgi/collection/all_epilepsy_seizuresAll Epilepsy/Seizuresfollowing collection(s): This article, along with others on similar topics, appears in the  Permissions & Licensing http://ng.neurology.org/misc/about.xhtml#permissionsits entirety can be found online at:Information about reproducing this article in parts (figures,tables) or in  Reprints http://ng.neurology.org/misc/addir.xhtml#reprintsusInformation about ordering reprints can be found online:reserved. 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De novo DNM1L mutation associated with mitochondrial epilepsy syndrome with fever sensitivity

De novo DNM1L mutation associated with mitochondrial epilepsy syndrome with fever sensitivity

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