Letters to the Editor. (bovine, mouse, rat ... suring the rates of ATP synthesis and electron transfer through ... of ATP synthesis to the rate of electron transfer. The.
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(bovine, mouse, rat, chicken, and Xenopus laevis) it is threonine, as in the LHON case studied here. Oxidative phosphorylation was studied in mitochondria from cultured lymphoblasts of patient III-1, an unaffected family member II-1, and two controls, by measuring the rates of ATP synthesis and electron transfer through complexes I and III simultaneously. In mitochondria carrying the ATPase 6/9101 mutation, the efficiency of oxidative phosphorylation was reduced by 40%-50%, as revealed by a lowered ATP/2e- ratio (III1, 0.75 + 0.07 [n = 3], and II-1, 0.78 + 0.06 [n = 3]; controls, 1.38 ± 0.14 [n = 7]), which relates the rate of ATP synthesis to the rate of electron transfer. The oligomycin-sensitive ATPase activity due to the FoF, complex was normalized to succinate dehydrogenase activity of the preparation, and a 20% decline was observed in the mutant preparation relative to that of the controls (III-1, 0.282 ± 0.025 [n = 3]; controls, 0.336 ± 0.055 [n = 3]). Two other mutations in ATPase subunit 6 have previously been associated with human disease. Heteroplasmic substitutions of Leu for Arg or Pro (de Vries et al. 1993) at residue 156 have been demonstrated in patients with either NARP (neurogenic muscle weakness, ataxia, and retinitis pigmentosa) or Leigh disease. We report now a new candidate for a primary mutation in LHON in a patient with typical symptoms of the disease. Although the site of the mutation is not in the conserved region of the ATPase subunit 6 gene, the biochemical defect in oxidative phosphorylation is specifically traced to complex V. To confirm the etiological role of the ATPase 6/9101 mutation in LHON, screening of the mutation is needed in LHON families from different populations. TARJA LAMMiNEN,1,2 ANNA MAJANDER,4 VESA JUVONEN,1 MARTEN WIKSTROM,4 PERTHr AULA,1 EEVA NIKOSKELAINEN,3 AND MARJA-LIISA SAVONTAUS1'2 Departments of 'Medical Genetics, 2Biology, and 3Ophthalmology, University of Turku, Turku, Finland; and 4Helsinki Bioenergetics Group, Institute of Biomedical Sciences, Department of Medical Chemistry, University of Helsinki, Helsinki
mutations associated with Leber's hereditary optic neuropathy. Genetics 130:163-173 de Vries DD, van Engelen BGM, Gabreels FJM, Ruitenbeek W. van Oost BA (1993) A second missense mutation in the mitochondrial ATPase 6 gene in Leigh's syndrome. Ann Neurol 34:410-412 Fillingame RH, Girvin ME, Fraga D, Zhang Y (1992) Correlations of structure and function in H' translocating subunit c of F1Fo ATP synthase. Ann N Y Acad Sci 671:323-333 Howell N, Bindoff LA, McCullough DA, Kubacka I, Poulton J, Mackey D, Taylor L, et al (1991) Leber hereditary optic neuropathy: identification of the same mitochondrial ND1 mutation in six pedigrees. Am J Hum Genet 49:939-950 Huoponen K, Lamminen T, Juvonen V, Aula P, Nikoskelainen EK, Savontaus M-L (1993) The spectrum of mitochondrial DNA mutations in families with Leber hereditary optic neuroretinopathy. Hum Genet 92:379-384 Huoponen K, Vilkki J, Aula P. Nikoskelainen EK, Savontaus M-L (1991) A new mtDNA mutation associated with Leber hereditary optic neuroretinopathy. Am J Hum Genet 48:1147-1153 Mackey D, Howell N (1992) A variant of Leber hereditary optic neuropathy characterized by recovery of vision and by an unusual mitochondrial genetic etiology. Am J Hum Genet 51:1218-1228 Marzuki S, Noer AS, Lertrit P, Thyagarajan D, Kapsa R, Utthanapol P, Byrne E (1991) Normal variants of human mitochondrial DNA and translation products: the building of a reference data base. Hum Genet 88:139-145 Vilkki J, Savontaus M-L, Nikoskelainen EK (1989) Genetic heterogeneity in Leber hereditary optic neuroretinopathy revealed by mitochondrial DNA polymorphism. Am J Hum Genet 45:206-211 (1990) Segregation of mitochondrial genomes in a heteroplasmic lineage with Leber hereditary optic neuroretinopathy. Am J Hum Genet 47:95-100 Wallace DC, Singh G, Lott MT, Hodge JA, Schurr TG, Lessa AMS, Elsas LJ II, et al (1988) Mitochondrial DNA mutation associated with Leber's hereditary optic neuroretinopathy. Science 242:1427-1430
Acknowledgments This work was supported by grants from the Sigrid Juselius Foundation, NIH (grant 1-RO1 EY09040-01), and the Academy of Finland (Medical Research Council). The technical assistance of Ilona Carlsson and Pirkko Jalava is gratefully acknowledged.
Exclusion of Chromosome 1q21 -q3 I from Linkage to Three Pedigrees Affected by the Pigment-
References Brown MD, Voljavec AS, Lott MT, Torroni A, Yang C-C, Wallace DC (1992) Mitochondrial DNA complex I and III
© 1995 by The American Society of Human Genetics. All rights reserved.
0002-9297/95/5605-0030$2.00
Am. J. Hum. Genet. 56:1240-1243, 1995
Dispersion Syndrome To the Editor: The pigment-dispersion syndrome is a form of openangle glaucoma that usually affects individuals in the first 3 decades of life. In addition to the typical opticnerve degeneration seen in all types of glaucoma, the pigment-dispersion syndrome is characterized by distinctive clinical features including the deposition of pig-
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ment granules from the iris epithelium on a variety of ocular structures including the trabecular meshwork. Frequently this disorder affects young myopic individuals. In the early stages of the disease, affected individuals may have clinical evidence of dispersed pigment without an associated elevation of intraocular pressure and optic-nerve degeneration. However, as the disease process progresses, many affected individuals (-50%) will develop elevated intraocular pressure and degeneration of the optic nerve, causing a permanent loss of sight (Richter et al. 1986). The pigment-dispersion syndrome shares several clinical features with the form of autosomal dominant juvenile open-angle glaucoma that recently has been mapped to the lq21-q31 region of chromosome 1 (Sheffield et al. 1993; Richards et al. 1994; Wiggs et al. 1994). Autosomal dominant inheritance has been described in a number of pedigrees affected by the pigment-dispersion syndrome (Becker and Podos 1966; Kaiser-Kupfer et al. 1983; Mandelkorn et al. 1983; McDermott et al. 1987). Both syndromes share a similar age at onset (the first 2 or 3 decades of life) and a high prevalence of myopia in affected individuals (Scheie and Cameron 1981; Johnson et al. 1993; J. L. Wiggs, unpublished data). Moreover, in our study of juvenile-glaucoma pedigrees, we identified an individual in one pedigree who was affected by the pigment-dispersion syndrome but who did not have elevated intraocular pressure or optic-nerve damage. Four of his five siblings, however, had severe juvenile glaucoma without any evidence of transillumination defects or other clinical features of the pigment-dispersion syndrome. We were subsequently able to determine that the juvenile glaucoma in this pedigree segregated with markers located on lq21-q31 and that the individual with pigment dispersion had inherited, from his affected father, the haplotype that did not segregate with the juvenile glaucoma, suggesting that, in this pedigree, the pigment-dispersion syndrome was genetically distinct from the form of juvenile glaucoma caused by the locus mapped to lq21-q31 (fig. 1; Wiggs et al. 1994). Phenotypic variability among individuals sharing a common gene defect has been observed in a number of human disorders, including such eye diseases as aniridia, retinitis pigmentosa, and forms of macular dystrophy (Berson et al. 1991; Nichols et al. 1993; Hanson et al. 1994). It is of interest, therefore, to investigate the spectrum of disease that may be generated by a gene known to be responsible for at least one form of juvenile glaucoma. To date, the only glaucoma pedigrees shown to be linked to the lq21-q31 region are those affected by a rare form of autosomal dominant juvenile glaucoma (Johnson et al. 1993; Sheffield et al. 1993; Richards et al. 1994; Wiggs et al 1994). To further investigate the genetic relationship between the pigment-dispersion syndrome and juvenile glaucoma, we have identified three pedigrees affected with the pigment-dispersion syn-
ADG-1S
7 D1S196
23
14
13
D1S445 3H D1S431 42 3H D1S210 D1S452 4 3H D1S242 D1S218 5 D1S416 I-
14 24 22 12 41 43 11
41
44
1 22 13 2 32 3
2
2
[ 3
4
134 112 1312
13L 4
33
1i1
53
14 31 1 232 3 233 31
1 33
43 24
14
4242 3
H
42 212 1 2[12 11 11 23 3
24 21 12 42 11 11 4 24 11
11 44 34 13 33 13 54 32
43 14
3
21 11 44
1434 4 13 22 33 23 1 1 31
43 31
51 32
41
14 13 22 23 31
41
32
Pigment dispersion syndrome
Juvenile glaucoma pedigree. Affected individuals are Figure I shown as blackened circles (females) or blackened squares (males). Deceased individuals not available for linkage analysis are indicated with slashes. Haplotypes are presented to illustrate the segregation of the affected parental chromosomes. The markers indicated in the pedigree drawing were used for this analysis. The alleles segregating with juvenile glaucoma are boxed. The probable location of recombination events is indicated by the square bracket. Individual ADG1812 is affected by the pigment-dispersion syndrome but not by juvenile glaucoma.
drome and have performed genetic linkage analysis using the collection of dinucleotide-repeat polymorphisms located on chromosome lq21-q31 that are positively linked to pedigrees affected with juvenile open-angle glaucoma. Twenty-two members of three pedigrees were identified for linkage analysis. Individuals were designated as affected for linkage purposes if all of the following findings typical of the pigment-dispersion syndrome were identified: pigment deposition on the corneal endothelium, iris transillumination defects, and increased pigmentation of the trabecular meshwork. Because we are investigating the genetic linkage of the pigment-dispersion syndrome, we did not require elevation of intraocular pressure or optic-nerve damage for affected status. However, 50% of the patients affected by the pigmentdispersion syndrome who were entered into the study had intraocular pressures >22, without medical or surgical treatment, and demonstrated associated opticnerve damage. The following microsatellites (Gyapay et al. 1994) were selected for segregation analysis (listed in proximal to distal order): D1S196-D1S433-D1S445-D1S4315 cM-DlS452-3 cM-D1S242-D1S218-D1S416 (the genetic distance between markers D1S196 and D1S431 and between D1S242 and D1S416 is -1 cM). Previous
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Table I Pairwise Linkage Data
Z AT O =
MARKER
A. Complete penetrance: D1S196........................... D1S433........................... DlS445........................... DlS431........................... DlS452........................... D1S242........................... DlS218........................... DlS416...........................
B. Affected individuals only: D1S196 ................... D1S433 ................... DlS445 ................... DlS431 ................... DlS452 ................... D1S242 ................... DlS218 ................... DlS416 ...................
.00
.01
.02
.03
.04
.05
.10
.20
-2 EXCLUSION
-00 -00 -00 -00
-7.00 -5.64 -6.47 -5.92 -8.05 -6.16 -7.73 -7.25
-5.40 -4.80 -5.21 -4.72 -6.41 -4.93 -6.21 -5.76
-4.66 -3.77 -4.93 -4.03 -5.45 -4.22 -5.34 -4.91
-4.07 -3.29 -3.91 -3.51 -4.75 -3.69 -4.68 -4.28
-3.60 -2.92 -3.50 -3.12 -4.21 -3.17 -4.19 -3.80
-2.22 -1.81 -2.42 -1.93 -2.79 -1.97 -2.67 -2.34
-.97 -.80 -1.61 -1.04 -1.25 -1.08 -1.39 -1.22
.11 .08 .10 .08 .12 .10 .14 .12
-00
-00 -00 -00
.00
.01
.015
.02
.025
.03
-9.07 -9.10 -4.40 -4.40 -9.08 -9.10 -9.10 -9.10
-2.80 -2.80 -1.40 -1.40 -2.80 -2.80 -2.80 -2.80
-2.46 -2.46 -1.23 -1.23 -2.46 -2.46 -2.46 -2.46
-2.22 -2.22 -1.11 -1.11 -2.22 -2.22 -2.22 -2.22
-2.02 -2.02 -1.01 -1.01 -2.02 -2.02 -2.02 -2.02
-1.86 -1.86 -.93 -.93 -1.86 -1.86 -1.86 -1.86
mapping studies have placed the critical region for juvenile glaucoma between D1S196 and DlS218, a distance of -8 cM (Sheffield et al. 1993; Richards et al. 1994; Wiggs et al. 1994). The six markers selected for this study comprehensively span the juvenile glaucoma critical region. Synthesis of oligonucleotide primers, amplification of microsatellites by PCR, and electrophoresis of the amplification products were performed as described elsewhere (Wiggs et al. 1994). Lod score (Z) values for pairwise analyses were calculated using the MLINK feature of the computer program LINKAGE, version 5.1 (Lathrop and Lalouel 1984). These calculations were performed using a 486/DX computer. The following assumptions were made: the spontaneous mutation rate was estimated to be 1 x 10-8, and the frequency of the mutant allele at the disease locus was estimated to be 1 x 106. Both are consensus figures from the literature, and alteration of these numbers had virtually no effect on the resulting Z values. The allele frequencies determined from studying 20 chromosomes of a similar patient population were comparable to those previously published, and hence the published allele frequencies were used in these calculations. Although formal segregation studies to estimate the penetrance of pigment dispersion have not been performed, approximately one-half (8/16) of the individuals at risk for the disease in the pedigrees presented here are affected, suggesting a penetrance close to 100%. Moreover, although the age at onset of this disorder
.025 .025 .005 .005 .025 .025 .025 .025
is characteristically before the 4th decade, occasionally cases are detected at older ages. In order to eliminate the risk of reduced penetrance due to delayed expression of the disease in an otherwise asymptomatic carrier, two models were assumed for linkage calculations, the first assuming a penetrance of 1.0 and the second using an affected-only analysis. Z(recombination fraction [0]) = -2.0 was used as evidence of exclusion. Pairwise linkage analyses exclude linkage of the pigment-dispersion trait to this collection of markers. The combined Z values from the three affected pedigrees provide evidence for exclusion of the entire 8-cM region (table 1). Haplotypes of affected and unaffected individuals from the three pedigrees studied are shown in figure 2. Nonsegregation of haplotypes derived by reconstruction with alleles from this collection of markers was observed for each of the three pedigrees. For example, two independent pairs of affected siblings (siblings PD13 and PD1-6 and siblings PD3-6 and PD3-8) inherited different copies of this region of chromosome 1 from the affected parent. Although the genetic heterogeneity of the pigment-dispersion syndrome is unknown, and although we cannot conclude that the same gene defect is responsible for the syndrome affecting these three pedigrees, we are able to exclude the lq21-q31 region in each pedigree, on the basis of the nonsegregation of the reconstructed haplotypes. Our results indicate that the pigment-dispersion syndrome, a form of glaucoma that may also affect the juvenile population, is genetically
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PD-1
tat D1S196 12 D1S433 33
[11 I 33
01S452 44
3
22
D1S218 41 133
018445 34 132I 13 II 12 2
PD-2018431
018242 22
122
D1S196
13
[32
01S431 12
13 22 12
3
3
2
018433 23 018445 12
32131i
33 12
018416 32
3
D1S452
44 D1S242 2 2
01S21842
Figure 2 fig. 1.
3
1
_3 a34
33
1
13
7
3
321
21 2
1
1
3 4
3
213
1
0
2j3
11
Pigment-dispersion pedigrees. Symbols are as in
unrelated to the autosomal dominant form of juvenile glaucoma caused by a defect in a gene located in the 1q21-q31 region of chromosome 1. CRISTINA PAGLINAUAN,1 JONATHAN L. HAINES,2 ELIZABETH A. DEL BONO,1 JOEL SCHUMAN,1 STEVEN STAWSKI,1 AND JANEY L. WIGGS1
tDepartment of Ophthalmology, New England Medical Center, Tufts University School of Medicine, and 2Neurogenetics Unit, Massachusetts General Hospital, Harvard Medical School, Boston
Berson EL, Rosier B, Sandberg MA, Dryja PT (1991) Ocular findings in patients with autosomal dominant retinitis pigmentosa and a rhodopsin gene defect (Pro-23-His). Arch Ophthalmol 109:92-101 Gyapay G, Morissette J, Vignal A, Dib C, Fizames C, Millasseau P. Marc S, et al (1994) The 1993-1994 Genethon human genetic linkage map. Nat Genet 7:246-339 Hanson IM, Fletcher JM, Jordon T, Brown A, Taylor D, Adams RJ, Punnett HH, et al (1994) Mutations at the PAX6 locus are found in heterogeneous anterior segment malformations including Peters' anomaly. Nat Genet 6:168-173 Johnson AT, Drack AV, Kwitek AE, Cannon RL, Stone EM, Alward WL (1993) Clinical feature and lineage analysis of a family with autosomal dominant juvenile glaucoma. Ophthalmology 100:524-529 Kaiser-Kupfer MI, Kupfer C, McCain L (1983) Asymmetric pigment dispersion syndrome. Trans Am Ophthalmol Soc 81:310-324 Lathrop GM, Lalouel JM (1984) Easy calculations of LOD scores and genetic risks on small computers. Am J Hum Genet 36:460-465 Mandelkorn RM, Hoffman ME, Olander KW, Zimerman T, Harsha D (1983) Inheritance and the pigmentary dispersion syndrome. Ann Ophthalmol 15:577-582 McDermott JA, Ritch R, Berger A, Wang RF (1987) Inheritance of pigmentary dispersion syndrome. Invest Ophthalmol Vis Sci Suppl 28:153 Nichols BE, Sheffield VC, Vandenburgh K, Drack AV, Kimura AE, Stone EM (1993) Butterfly-shaped pigment dystrophy of the fovea caused by a point mutation in codon 167 of the RDS gene. Nat Genet 3:202-207 Richards JE, Lichter PR, Boehnke M, Uro JLA, Torrez D, Wong D, Johnson AT (1994) Mapping of a gene for autosomal dominant juvenile-onset open-angle glaucoma to chromosome lq. Am J Hum Genet 54:62-70 Richter CU, Richardson TM, Grant WM (1986) Pigmentary dispersion syndrome and pigmentary glaucomas: a prospective study of the natural history. Arch Ophthalmol 105:211-215 Scheie HG, Cameron JD (1981) Pigment dispersion syndrome: a clinical study. Br J Ophthalmol 65:264-269 Sheffield, VC, Stone EM, Alward WLM, Drack AV, Johnson AT, Streb LM, Nichols BE (1993) Genetic linkage of familial open angle glaucoma to chromosome lq21-q31. Nat Genet 4:47-50 Wiggs JL, Haines JL, Paglinauan C, Fine A, Sporn C, Lou D (1994) Genetic linkage of autosomal dominant juvenile glaucoma to 1q21-q31 in three affected pedigrees. Genomics 21:299-303
Acknowledgments
© 1995 by The American Society of Human Genetics. All rights reserved. 0002-9297/95/5605-0031$2.00
We would like to thank the families for their participation in this study; Derek Lou, Madalina Pralea, and Catalin Pralea for their contributions to the study; and Cheryl Connery for administrative assistance. This work was supported in part by NIH grant EY09847 to J.L.W.
Am. J. Hum. Genet. 56:1243-1245, 1995
References
Molecular Analysis of Glucose-6-Phosphate Dehydrogenase Variants in the Solomon Islands To the Editor:
Becker B, Podos SM (1966) Krukenberg's spindles and primary open-angle glaucoma. Arch Ophthalmol 76:635-639
is one of the most prevalent genetic disorders, and > 100
Glucose-6-phosphate dehydrogenase (G6PD) deficiency
