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Leigh Syndrome Associated with Mitochondrial Complex I Deficiency Due to a Novel Mutation in the NDUFS1 Gene

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Leigh Syndrome Associated with Mitochondrial Complex I Deficiency Due to a Novel Mutation in the NDUFS1 Gene OBSERVATION Leigh Syndrome Associated With Mitochondrial Complex I Deficiency Due to a Novel Mutation in the NDUFS1 Gene Miguel A. Martín, PhD; Alberto Blázquez, BSc; Luis G. Gutierrez-Solana, MD; Daniel Fernández-Moreira, PharmB; Paz Briones, PhD; Antoni L. Andreu, MD, PhD; Rafael Garesse, PhD; Yolanda Campos, PhD; Joaquín Arenas, PhD Background: Mutations in the nuclear-encoded sub- Results: Muscle biochemistry results showed a com- units of complex I of the mitochondrial respiratory chain plex I defect of the mitochondrial respiratory chain. Se- are a recognized cause of Leigh syndrome (LS). Re- quencing analysis of the mitochondrial DNA–encoded cently, 6 mutations in the NDUFS1 gene were identified ND genes, the nuclear DNA–encoded NDUFV1, NDUFS1, in 3 families. NDUFS2, NDUFS4, NDUFS6, NDUFS7, NDUFS8, and NDUFAB1 genes, and the complex I assembly factor CIA30 Objective: To describe a Spanish family with LS, com- gene revealed a novel homozygous L231V mutation plex I deficiency in muscle, and a novel mutation in the (c.691C→G) in the NDUFS1 gene. The parents were het- NDUFS1 gene. erozygous carriers of the L231V mutation. Design: Using molecular genetic approaches, we iden- Conclusions: Identifying nuclear mutations as a cause tified the underlying molecular defect in a patient with of respiratory chain disorders will enhance the possibil- LS with a complex I defect. ity of prenatal diagnosis and help us understand how mo- lecular defects can lead to complex I deficiency. Patient: The proband was a child who displayed the clini- cal features of LS. Arch Neurol. 2005;62:659-661 EIGH SYNDROME (LS) (ON- in the NDUFS1 gene were identified in 3 line Mendelian Inheritance families with LS or Leigh-like disease and in Man 256000) is a devas- complex I deficiency (Online Mendelian In- tating neurodegenerative heritance in Man 157655).5 Herein, we de- disorder characterized neu- scribe a Spanish patient with LS, complex Lropathologically by focal bilaterally sym- I deficiency in muscle, and a novel muta- metrical lesions, especially in the thala- tion in the NDUFS1 gene. mus and brainstem regions, and clinically by psychomotor retardation, respiratory dif- METHODS ficulties, nystagmus, ophthalmoparesis, op- tic atrophy, ataxia, and dystonia.1 In most patients, mitochondrial respiratory chain REPORT OF A CASE defects and pyruvate dehydrogenase com- The first child (a girl) of healthy nonconsan- plex deficiency are the underlying causes 2 guineous parents (aged 30 years) of Spanish of the disease. Mitochondrial respiratory origin was born after a term pregnancy (birth chain complex I (nicotinamide adenine di- weight, 3560 g). She was hospitalized at age 8½ nucleotide:ubiquinone oxidoreductase) months for recurrent episodes of vomiting, contains at least 46 subunits, 7 of which are floppiness, and growth retardation. She pre- encoded by mitochondrial DNA (mtDNA).2 sented with irritability, horizontal nystagmus, Various mutations in a few subunits of com- and generalized hypotonia. Tendon reflexes plex I encoded by nuclear DNA (nDNA) were hyperactive and symmetric. Babinski sign (NDUFV1, NDUFV2, NDUFS1, NDUFS2, was negative. An electroencephalogram dis- NDUFS3, NDUFS4, NDUFS7, and played a normal pattern. Brain magnetic reso- 3-10 nance imaging showed bilateral lesions affect- NDUFS8) are associated with LS or ing the substantia nigra and midbrain. Leigh-like disease in patients with com- Cardiologic and ophthalmologic examination plex I deficiency. The NDUFS1 gene en- findings were normal. Laboratory data revealed Author Affiliations are listed at codes the largest (75-kDa subunit) pro- increased lactate levels in blood (24 mg/dL; the end of this article. tein of complex I.11 Recently, 6 mutations normal, Ͻ20 mg/dL) and in cerebrospinal fluid (REPRINTED) ARCH NEUROL / VOL 62, APR 2005 WWW.ARCHNEUROL.COM 659 ©2005 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 10/02/2021 A 1234 B T A GGT GCCCT AA CCT Table. Activities of Mitochondrial Respiratory Chain ∗ Complexes in Muscle Homogenate* 312 bp 180 bp 132 bp Control Subjects, TTTAGGT GGCCAA CC Mean (SD) ∗ Activity Patient (n = 100) Nicotinamide adenine dinucleotide: 5.0 20.0 (4.5) ubiquinone oxidoreductase L231V C Succinate dehydrogenase 8.7 10.3 (3.5) Homo sapiens SGN I ID I CPVGALTSKPYAFTARPWETRKTESIDVMDAVG Decylubiquinol–cytochrome c oxidoreductase 50.7 63.0 (18.0) Bos taurus SGN I ID I CPVGALTSKPYAFTARPWETRKTESIDVMDAVG Cytochrome c oxidase 22.8 42.0 (12.1) Solanum tuberosum SGN V ID I CPVGALTSKPFAFKARNWELKGTESIDV T DAVG Rhodobacter capsulatus QGN I IDLCPVGALTSKPYAFTARPWELVKTESIDVMDALG *Data are given as percentage of citrate synthase activity. Activities were Neurospora crassa SGN V IDLCPVGALTSKPYAFRARPWELKKTESIDV L DGLG measured as nanomoles per minute per milligram of protein. Paracoccus denetrificans QGN I IDLCPVGALVSKPYAFTARPWELT KTESIDVMDALG Rickettsia prowazekii SGN MID I CPVGALNSKPYAFKARKWELKHTASIGV H DAEG Figure. Polymerase chain reaction–restriction fragment length polymorphism results (A), sequencing (B), and sequence alignment of the RESULTS NDUFS1 protein from various species (C). A, Lane 1: control DNA. Lane 2: muscle DNA from the patient. Lane 3: blood DNA from the mother. Lane 4: blood DNA from the father. The L231V mutation was detected by digesting The activities of respiratory chain complexes in muscle the NDUFS1 exon 8 fragment with the restriction enzyme Sdu I. The showed a single defect of nicotinamide adenine dinucleo- 312–base pair (bp) wild-type DNA was digested by Sdu I into 2 fragments of tide:ubiquinone oxidoreductase (complex I), account- 180 and 132 bp, whereas the 312-bp fragment remained uncut in the patient, Table who was homozygous for the mutation. B, Electrophoretograms showing the ing for 25% of the mean of the control subjects ( ). change in the patient (top) and the normal sequence in a control subject Given the clinical picture and the biochemical findings, (bottom). Asterisk represents the nucleotide substitution. C, Evolutionary we searched for the underlying molecular alteration of conservation of the 231 amino acid residue (arrow) of the NDUFS1 protein. this defect. Sequencing analysis of the genes listed in the “Methods” section showed a novel homozygous mis- (30 mg/dL; normal, Ͻ15 mg/dL). In fibroblasts, pyruvate dehy- sense mutation (L231V) that replaces a leucine by a va- drogenase activity was normal, and pyruvate oxidation was in the line in the 231 amino acid residue of the protein as a re- lower reference limit. Muscle morphologic structure was nor- sult of a c.691C→G transversion in exon 8 of the NDUFS1 mal except for mild atrophy of type 2 fibers. Her status wors- gene (Figure). Additional nucleotide changes were not ened 5 months later, when she developed respiratory insuffi- found. The parents were heterozygous carriers of the ciency and horizontal and vertical nystagmus. Her tendon reflexes L231V mutation. became highly hyperactive, with a surface area enlargement and presence of Babinski sign. She died at age 14 months. Her younger brother had a similar clinical picture and died at age 8 months, COMMENT after an acute episode of respiratory failure. Mutations in the nDNA-encoded complex I NDUFV1, BIOCHEMICAL AND MOLECULAR NDUFV2, NDUFS1, NDUFS2, NDUFS3, NDUFS4, GENETIC STUDIES NDUFS7, and NDUFS8 genes have been documented in patients with LS, although often the underlying molecu- An appropriate institutional review board approved this work, lar defect remains unknown.3-10 In 3 nonrelated fami- and informed consent was obtained from the child’s parents. lies, Bénit et al5 identified 6 mutations in the NDUFS1 Respiratory chain enzymes in muscle homogenate were mea- gene, which encodes the largest subunit of complex I. sured by methods reported elsewhere.12 Interestingly, 3 of these mutations (amino acids 222, 241, DNA was isolated from muscle and blood from the patient and 252) lie in a highly evolutionary conserved stretch and from blood from her parents. The mtDNA-encoded ND sub- 13 of the protein encompassing the most C-terminal cyste- units were amplified using suitable primers. The coding re- ine residue, potentially involved in the ligation of iron- gion and exon and intron boundaries of the nuclear-encoded 5 complex I subunits of the NDUFV1, NDUFS1, NDUFS2, sulfur clusters. NDUFS3, NDUFS4, NDUFS7, NDUFS8, and NDUFAB1 genes, We describe a Spanish girl with LS, whose younger as well as the complex I assembly factor CIA30 gene, were am- brother died of a similar condition. The proband had a plified as previously described3-10,14,15 or by using novel in- complex I defect in muscle and harbored a novel homo- tronic primers. Polymerase chain reaction products were pu- zygous missense mutation (L231V) in the NDUFS1 gene. rified by electrophoresis in 2% agarose gel and sequenced The mutation was consistently heterozygous in blood directly, using the ABI PRISM dRhodamine Terminator Cycle DNA from the healthy parents, suggesting autosomal re- Sequencing Kit in an ABI PRISM 310 genetic analyzer (Ap- cessive inheritance. Several lines of evidence support the plied Biosystems, Foster City, Calif). Nucleotide changes were pathogenicity of the mutation, including the following: further confirmed by polymerase chain reaction–restriction frag- (1) the patient had a single complex I defect in muscle; ment length polymorphism methods (Figure). One hundred healthy control subjects (200 alleles) were screened by poly- (2) it was the only nucleotide change found in the en- merase chain reaction–restriction fragment length polymor- tire coding region and in the intron and exon bound- phism methods to rule out the presence of the mutation in the aries of the gene; (3) no additional pathogenic muta- healthy population. Unfortunately, tissue specimens were not tions were found in the other complex I genes analyzed; available from the proband’s brother. (4) the mutation was absent in 200 alleles from 100 (REPRINTED) ARCH NEUROL / VOL 62, APR 2005 WWW.ARCHNEUROL.COM 660 ©2005 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 10/02/2021 healthy controls of similar ethnic background; and (5) Fernández-Moreira, and Arenas.
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