Cytochrome C Oxidase Biogenesis in a Patient with a Mutation in COX10
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ampullopetal deflection and inhibition of the superior canal.1,9 Clockwise direction of the slow phase indi- Cytochrome c Oxidase cates predominant inhibition of the right superior ca- Biogenesis in a Patient with nal that might be more affected than the left one. The permanent oscillopsia in our patient might be a Mutation in COX10 Gene explained by abnormal visual inhibition of VOR, Marieke J. H. Coenen, MSc,1 which in association with gaze-evoked nystagmus and Lambert P. van den Heuvel, PhD,1 Cristina Ugalde, PhD,1 the abnormal clinical aspect of smooth pursuit might Marike ten Brinke,1 Leo G. J. Nijtmans, PhD,1 be linked to cerebellar lithium toxicity. Indeed, these Frans J. M. Trijbels, PhD,1 Skadi Beblo, MD,2 Esther M. Maier, MD,2 Ania C. Muntau, MD,2 different cerebellar oculomotor symptoms are known 1 to occur with lithium therapy even within the range of and Jan A. M. Smeitink, MD, PhD that drug’s therapeutic blood level.11 In conclusion, this article adds important data to the We report a cytochrome c oxidase (COX)–deficient pa- clinical description and pathophysiological understand- tient, clinically affected with Leigh-like disease, with a ing of superior canal dehiscence syndrome. Vertical os- homozygous mutation in the COX10 start codon. Two- cillopsia and pulse-synchronous nystagmus may be ob- dimensional gel electrophoresis showed a decrease of served in bilateral symptomatic forms as a result of an fully assembled COX without the accumulation of par- abnormal communication between the inner ear and tially assembled COX subcomplexes. Western blot analy- intracranial space. sis with antibodies directed to COX subunits I, II, and IV showed a decrease of these subunits in this patient compared with control. Overexpression of the COX10 We thank Dr G. Rambaud for referring the patient described in this protein in the patient’s fibroblasts proved that the de- article. tected mutation was indeed the disease cause. References Ann Neurol 2004;56:560–564 1. Minor LB, Solomon D, Zinreich JS, Zee DS. Sound- and/or pressure-induced vertigo due to bone dehiscence of the superior semicircular canal. Arch Otolaryngol Head Neck Surg 1998; Human cytochrome c oxidase (COX) consists of 13 124:249–258. subunits; three of these are encoded by the mitochon- 2. Carey JP, Minor LB, Nager GT. Dehiscence or thinning of drial DNA. Because of the bigenomic origin of the bone overlying the superior semicircular canal in a temporal complex, isolated COX deficiencies can be caused by bone survey. Arch Otolaryngol Head Neck Surg 2000;126: 137–147. mutations in either the mitochondrial or the nuclear 3. Younge BR, Khabie N, Brey RH, Driscoll CL. Rotatory nys- genome. In contrast to complexes I, II, and III, no tagmus synchronous with heartbeat: a treatable form of nystag- mutations have yet been described in any nuclear- mus. Trans Am Ophthalmol Soc 2003;101:113–117; discus- encoded structural subunit of COX.1–3 However, six sion 117–118. genes involved in COX biogenesis have been linked to 4. Brantberg K, Bergenius J, Mendel L, et al. Symptoms, findings and treatment in patients with dehiscence of the superior semi- COX deficiency in humans (SURF1, SCO1 and SCO2, 4–13 circular canal. Acta Otolaryngol 2001;121:68–75. COX10, COX15, and LRPPRC). The COX10 and 5. Cremer PD, Minor LB, Carey JP, Della Santina CC. Eye COX15 proteins play a role in the mitochondrial heme movements in patients with superior canal dehiscence syndrome biosynthetic pathway. COX10 catalyzes the conversion align with the abnormal canal. Neurology 2000;55:1833–1841. of protoheme to heme O. COX15 exerts its role in the 6. Mong A, Loevner LA, Solomon D, Bigelow DC. Sound- and pressure-induced vertigo associated with dehiscence of the roof next step, in which heme O is converted to heme A, an 10 of the superior semicircular canal. AJNR Am J Neuroradiol essential group for the functioning of complex IV. 1999;20:1973–1975. To date, three patients harboring mutations in COX10 7. Deutschlander A, Strupp M, Jahn K, et al. Vertical oscillopsia in bilateral superior canal dehiscence syndrome. Neurology 2004;62:784–787. 8. Rambold H, Heide W, Sprenger A, et al. Perilymph fistula as- From the 1Department of Paediatrics, Nijmegen Centre for Mito- sociated with pulse-synchronous eye oscillations. Neurology chondrial Disorders, University Medical Centre Nijmegen, Nijme- 2001;56:1769–1771. gen, The Netherlands; and 2Metabolic Department, Dr. von 9. Hirvonen TP, Carey JP, Liang CJ, Minor LB. Superior canal Hauner Children’s Hospital, Ludwig-Maximilians-University, Mu- dehiscence: mechanisms of pressure sensitivity in a chinchilla nich, Germany. model. Arch Otolaryngol Head Neck Surg 2001;127: Received Apr 15, 2004, and in revised form Jun 14. Accepted for 1331–1336. publication Jun 15, 2004. 10. Leigh RJ, Zee DS. The neurology of eye movements. 3rd ed. Published online Sep 30, 2004 in Wiley InterScience Philadelphia: F. A. Davis, 1999. (www.interscience.wiley.com). DOI: 10.1002/ana.20229 11. Corbett JJ, Jacobson DM, Thompson HS, et al. Downbeating nystagmus and other ocular motor defects caused by lithium Address correspondence to Dr Van den Heuvel, Department of Pae- toxicity. Neurology 1989;39:481–487. diatrics, University Medical Centre Nijmegen, 6500 HB Nijmegen, The Netherlands. E-mail: [email protected] 560 © 2004 American Neurological Association Published by Wiley-Liss, Inc., through Wiley Subscription Services Fig 1. Molecular analysis of COX10 genomic DNA. (A) COX10 DNA sequence of control and patient blood; the arrow indicates the mutation in the start codon of COX10. (B) Restriction endonuclease analysis pattern with BstZI. The fragment harboring the mutation could be digested, whereas the control was undigested. The length of the fragments (in base pairs) are indicated on the right site of the figure. p, patient; f, father; m, mother; w, wild type. have been described.8,9 Here we report a new patient, tivities were measured in skin fibroblasts and muscle (slightly phenotypically classified as suffering from a Leigh-like modified from the method described previously14). disease, with a mutation in the start codon of the COX10 gene. The effect of this mutation on the as- Analysis of COX10 DNA sembly or stability of mitochondrial COX has been an- A group of 11 patients with an isolated COX deficiency at alyzed by two-dimensional blue-native electrophoresis. least expressed in cultured skin fibroblasts were included in this study. DNA was isolated from patients’ fibroblasts and parents’ blood.15 The oligonucleotide primers used for the Case Report amplification of COX10 genomic DNA were described pre- The male patient was born at term as the first child of viously.8 The DNA sequence was analyzed on an ABI 377 consanguineous parents. At 5 months’ age, he devel- sequencer (Perkin-Elmer, Oak Brook, IL). To confirm the oped progressive failure to thrive, and pronounced mo- presence of the mutation, we performed restriction fragment tor agitation was noted. Gross motor development was length polymorphism analysis with BstZI (Promega, Madi- severely delayed at 7 months. At this age, the patient sion, WI). showed generalized muscular hypotonia with persistent head lag at traction, ataxia, hypermetria, exaggerated COX10 Complementary DNA Construct, Virus tendon reflexes with enlarged reflex zones, low- Production, Infection, and Measurement amplitude nystagmus, and saccadic eye movements. of Enzyme Activity 9 Ocular fixation was weak. He was not able to grasp. The retroviral vector was created as described previously. Laboratory evaluation showed metabolic acidosis with COX activities were measured before and after overexpres- sion of COX10 protein (as described by Capaldi and col- elevated serum and cerebrospinal fluid lactate concentra- leagues16 and Srere17). tions. Magnetic resonance imaging of the brain showed slight atrophy and hyperintense lesions in the thalamus, Protein Electrophoresis olives, and the nucleus ruber, a pattern comparable to One- and two-dimensional blue-native electrophoresis were Leigh-like disease. Biochemical COX activity was signif- performed with digitonin-isolated mitochondria.18 Sodium icantly reduced in muscle and fibroblasts (0.15 COX/ dodecyl sulfate polyacrylamide gel electrophoresis (SDS- citrate synthase [CS]; control range, 0.52–2.08; and PAGE) was performed according to the method of Schagger 0.22 COX/CS; control range, 0.68–1.19 COX/CS, re- and von Jagow.19 Proteins were transferred to a PROTAN spectively). The boy died at 9 months of age of acute nitrocellulose membrane (Schleicher & Schnell, Keene, NH). pneumonia and cardiorespiratory failure. Prenatal diag- Western blotting was performed using anti-COXI, anti- nosis was performed in a later pregnancy. Normal COX COXII, anti-COXIV (all from Molecular Probes, Eugene, activity was found in chorionic villi, and the mother OR), anti–mitochondrial HSP70 (Alexis, Molecular Probe, Eugene, OR), and peroxidase-conjugated anti–mouse immu- gave birth to a healthy girl. noglobulin G (Molecular Probes). The signal was detected by enhanced chemiluminescence with ECL Plus (Amersham Materials and Methods Biosciences, Arlington Heights, IL). Cell Culture and Biochemical Measurements Human skin fibroblasts were cultured in M199 (Life Tech- Results nologies, Bethesda, MD) supplemented with 10% fetal calf Eleven patients with decreased COX activity estab- serum and antibiotics. Mitochondrial OXPHOS complex ac- lished in cultured fibroblasts (data not shown) were an- Coenen et al: Novel COX10 Gene and COX Assembly 561 alyzed for mutations in the COX10 gene. A homozy- tection of COX subcomplexes. The patient’s fibroblasts gous mutation in the COX10 start codon was detected display a general decrease of all subcomplexes as well as in one patient. The T3C transition of the second a lesser amount of holo COX (Fig 2B). This general base of the start codon (ATG) results in the abolition decrease of the subcomplexes was also detected with an of the start site for protein translation (Fig 1A).