|Molecular Vision 2006;
Received 8 July 2006 | Accepted 19 July 2006 | Published 24 July 2006
Peripherin/RDS and VMD2 mutations in macular dystrophies with adult-onset vitelliform lesion
Stanislav A. Zhuk,1
Albert O. Edwards1-3
1Department of Ophthalmology and 2McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, TX; 3Institute for Retina Research, Dallas, TX
Correspondence to: Albert O. Edwards, MD, PhD, Institute for Retina Research, 8210 Walnut Hill Lane, Suite 010, Dallas, TX, 75231; Phone: (214) 345-6801; FAX: (214) 987-1881; email: email@example.com
Purpose: Adult-onset vitelliform macular dystrophy (AVMD) is a pleomorphic late-onset macular phenotype characterized by a central yellow deposit between the neural retina and retinal pigment epithelium. Mutations in the genes encoding peripherin/RDS and VMD2 have been previously reported in some subjects with AVMD. The purpose of this investigation was to determine the prevalence of mutations in these two genes in a cohort of cases with macular dystrophies presenting with vitelliform lesions in adulthood.
Methods: Fifty nine consecutively ascertained and unrelated subjects prospectively coded as pattern or vitelliform macular dystrophies were reviewed and twelve subjects with a vitelliform lesion were identified. Patient evaluation included comprehensive ocular examination, retinal imaging, and functional studies in selected subjects. The RDS and VMD2 genes were screened for variation by direct DNA sequencing of coding regions and intron/exon boundaries.
Results: Twenty-two DNA sequence variants were identified in the genes encoding RDS and VMD2. A Pro210Arg variant found in the RDS gene of one subject was the only definite mutation detected in either gene.
Conclusions: The Pro210Arg mutation has been reported previously in patients with pattern dystrophy confirming the observation that pattern dystrophy can present with an AVMD phenotype. Although RDS and VMD2 are the only known genes with mutations contributing to AVMD, our series demonstrates that most patients have mutations in genes that have yet to be discovered.
Adult-onset vitelliform macular dystrophy (AVMD) was described by Gass in 1974 . It presents with funduscopic findings overlapping Best disease . The primary feature is a slightly elevated, yellow, egg yolk-like lesion located in the foveal or parafoveal region. The lesion is one-third to one-half of disk diameter in size, either round or slightly oval, and with an occasional pigment spot in the center. The clinical presentation of AVMD, however, differs from Best macular dystrophy. The onset occurs later in life, usually after the age of 30, and presenting symptoms include blurred vision and metamorphopsia. The progression of visual loss is slow and most patients maintain reading vision in at least one eye in spite of later degenerative changes [2,3]. The vitelliform lesion typically disappears later in life. The electrooculogram is usually normal or subnormal, while the lectroretinogram and color vision are typically normal.
The genes encoding RDS and VMD2 have been implicated in some cases of AVMD. To date, 9 different mutations spanning all three exons of the RDS gene have been documented in AVMD patients [4-8]. VMD2 mutations were detected in approximately 96% of patients with Best macular dystrophy with a family history . Five different VMD2 mutations have also been found in patients with AVMD [10-12]. Two of these mutations have been reported in families with Best disease. The genetic heterogeneity underlying macular dystrophies with adult-onset vitelliform phenotype is largely unknown.
The goal of this study was to determine the prevalence of mutations in the genes encoding VMD2 and RDS in pathogenesis of macular dystrophies with onset of vitelliform lesions in adulthood. Entire coding regions of these two genes were screened for mutations in 12 unrelated patients diagnosed with AVMD. We show that mutations in VMD2 and RDS are uncommon causes of AVMD.
The study was approved by the institutional review board at University of Texas Southwestern Medical Center and all subjects gave written consent for participation. The subjects were prospectively ascertained. Patient evaluation included ocular history, best corrected visual acuity, slit-lamp biomicroscopy, dilated fundus examination, and fundus photography and/or optical coherence tomography. Fluorescein angiography and functional evaluations were performed as indicated for patient management. Family members related to subject 2 were examined.
The diagnosis of an adult-onset macular dystrophy with vitelliform lesion was established upon the presence of the following: (1) elevated yellow lesion between the neurosensory retina and retinal pigment epithelium by biomicroscopy and stereoscopic fundus photography in the foveal or parafoveal region of at least one eye; and (2) the onset of the dystrophy after the fourth decade. Fundus photographs from subjects with vitelliform lesions were also examined for other features previously reported in macular dystrophies with the vitelliform presentation: geographic atrophy, drusen, hyperpigmentation around the lesion, and choroidal neovascularization. Subjects with known Best macular dystrophy were excluded based on typical presentation, family history, or known mutation.
Blood samples were obtained by peripheral venipuncture and genomic DNA purified using standard methods . Primer pairs for DNA amplification were designed based on the genomic sequences of RDS and VMD2; a minimum of 50 base pairs flanking each exon were included in the amplification product. Polymerase chain reaction conditions and primers are available upon request. Amplification DNA fragments were verified for appropriate length and absence of nonspecific amplification products by agarose gel electophoresis. Amplified DNA was purified (QIAquick, Qiagen, Valencia, CA) and directly sequenced using ABI Dye Terminator 3.1 chemistry with ABI capillary instruments (Applied Biosystems, Foster City, CA). Mutations were defined as a heterozygous nucleotide change or insertion/deletion that resulted in a change in the peptide sequence or disruption of a splice acceptor or donor sequence not found in normal subjects. Known polymorphisms were identified using the Retina Mutation Database International, the VMD2 database of the Institute of Human Genetics, University of Regensburg, Germany, and the National Center for Biotechnology Information, Bethesda, MD.
Identification of subjects with adult-onset vitelliform macular dystrophy
A total of 59 consecutively ascertained subjects initially diagnosed with an adult onset macular dystrophy (excluding Best disease and age-related macular degeneration) were reviewed. Twelve were identified meeting the diagnostic vitelliform criteria of the study. The clinical data of the selected patients are summarized in Table 1. One patient was of African descent (Case 4) and the rest were Caucasian.
Clinical features of adult-onset vitelliform macular dystrophy subjects
The age at the time of diagnosis ranged from 48 to 86 years and the average age was 69.7 years. Five patients (42%) had a family history of macular dystrophy or degeneration. The visual acuity ranged from 20/20 to 20/70, excluding one subject with other reasons for vision loss. Hyperpigmentation was present in 8 patients (67%); drusen or flecks were found in 5 patients (42%); and atrophy of retinal pigment epithelium was found in one patient (8%). Choroidal neovascularization was not associated with any of the vitelliform lesions in this study. None of the families with AVMD had a history of vitreoretinal degeneration, nanophthalmus, or other developmental eye disorder.
Direct sequencing of PCR products spanning all 3 exons of the RDS/peripherin gene revealed a Pro210Arg mutation in subject 2 (Table 2). No other sequence variants in this gene were felt to be mutations (Table 2). No sequence variants in the 11 exons of the VMD2 gene changed the amino acid sequence of the VMD2 gene or the splice donor and acceptor sites (Table 3).
Clinical features associated with the Pro210Arg mutation
The patient with the Pro210Arg mutation was a 50 year-old Caucasian woman (subject 2) who presented after her brother was diagnosed with AVMD. The best-corrected visual acuity was 20/70 OD and 20/25 OS. Her fundus photographs revealed characteristic vitelliform lesions in the macula of both eyes (Figure 1). Her electroretinogram was normal; an electro-oculogram was not performed. Fundus photographs of her mother showed geographic atrophy of the macula bilaterally.
The discovery of the genetic basis of early-onset macular dystrophies facilitated their clinical classification and helped clarify the genotype-phenotype relationships [8,13-19]. However, the relationship between genotype and phenotype for the middle-age and late-age onset pattern, vitelliform, and other macular dystrophies remains poorly understood. AVMD falls into this category of middle-age and late-age onset macular dystrophies and is of considerable interest given the overlap in age of presentation and fundus features such as drusen, vitelliform lesions, and choroidal neovascularization. Our study confirms that the only two known genes (RDS and VMD2) implicated in AVMD are involved in a small proportion of cases [10-12]. Further, the genetic basis of most subjects with pattern dystrophy remains unknown. Thus, the genetic causes of the two most common groups of adult-onset macular dystrophies remain largely unknown.
The pattern dystrophies and AVMD are often asymptomatic at the initial stages when the characteristic lesions (e.g., vitelliform maculopathy) are visible. The characteristic lesions may disappear over time as atrophy of the outer retina progresses. Thus, the same patient at different times might be classified as AVMD, pattern dystrophy, and eventually atrophic age-related macular degeneration. To circumvent this difficulty in classification, we required the presence of a vitelliform lesion in our diagnostic criteria. It is probable that some of the other 47 subjects with macular dystrophies we reviewed had or will develop a vitelliform lesion.
We also report a fifth family segregating a Pro210Arg mutation in the RDS gene. Two siblings were found to have AVMD (subject 2 and her younger brother by six years). Their mother had extensive bilateral geographic atrophy of the macula. The first report of a Pro210Arg mutation was a case study of a patient who was diagnosed with AVMD and over the next twenty months developed choroidal neovascularization in one of eyes . The second study described three unrelated families with a Pro210Arg mutation, one of which was initially described by Gass in his original 1974 paper on AVMD [1,6]. These three families exhibited intrafamilial variability of phenotypes and genetic expressivity. The clinically affected members ranged in age from 16 to 75 years, visual acuities ranged from 20/20 to less than 20/200, and perimetric findings included severe peripheral vision loss, general threshold elevation, and central or paracentral scotomas. Retinal findings of the largest family were diverse and consisted of hard drusen, pattern dystrophy, vitelliform lesions (in 3 family members out of 12), extensive geographical atrophy, severe atrophic macular degeneration, and diffuse pigment epithelial disturbances.
The Pro210Arg mutation replaces the hydrophobic ring of proline with the positively-charged, large-branched amino acid arginine. Amino acid 210 is located in a large intradiscal loop that connects the third and fourth transmembrane segments. This loop contains seven highly conserved cysteine residues that may contribute to protein folding; it has been implicated in mediation of RDS/peripherin and ROM-1 protein interaction as well as in membrane fusion [20-22]. Interestingly, Kemp and coworkers reported a patient with autosomal dominant retinitis pigmentosa who had a point mutation at the same amino acid, but the substituted residue was hydrophilic serine . The abnormal function may be the result of a misfolded loop, different configurations of which can cause quite different retinal diseases. A majority of disease-causing mutations are located in this loop.
Although this study did not find any mutations in the VMD2 gene, two previous studies have found 5 different missense mutations associated with AVMD [11,12]. The investigators of both studies raised the possibility that these patients represent atypical cases of Best disease. First, two of the mutations, Ala243Val and Thr6Pro, have been previously found in patients with Best vitelliform dystrophy . Secondly, EOG, with values typically abnormal in Best disease and therefore useful in distinguishing it from AVMD, is not 100% specific. For example, Best disease patients with Ala243Val mutation seem to have EOG values either normal or higher than typically seen with the disorder . In the paper by Seddon et al., the AVMD patient with Ala146Lys mutation had a family history consistent with autosomal dominant pattern and EOG values in the range of Best disease . These VMD2 mutations may cause less severe forms of Best disease with later onset. Milder phenotype with near-normal EOGs and reduced penetrance that obscures family history could lead to diagnosis of these patients as having AVMD.
Vitelliform lesions do not appear to represent a phenotype limited to mutations in specific genes. Rather, the vitelliform lesion is a retinal response to disease observed with variable frequency in a number of conditions ranging from early-onset Best disease to age-related macular degeneration . Prior to this study, over 35 subjects with AVMD had been screened for mutations in VMD2, with 9 demonstrating 5 different mutations and over 30 subjects with AVMD had been screened for mutations in RDS/peripherin, with 5 families and 6 unrelated individuals demonstrating 7 different mutations. Thus, mutations in these two genes account for a minority of AVMD cases, an observation further supported by our study [10-12].
We thank Mr. Robert Ritter and the McDermott Sequencing Center at UTSW for assistance. Supported by the National Eye Institute (EY014467) and the Foundation Fighting Blindness.
1. Gass JD. A clinicopathologic study of a peculiar foveomacular dystrophy. Trans Am Ophthalmol Soc 1974; 72:139-56.
2. Patrinely JR, Lewis RA, Font RL. Foveomacular vitelliform dystrophy, adult type. A clinicopathologic study including electron microscopic observations. Ophthalmology 1985; 92:1712-8.
3. Burgess DB, Olk RJ, Uniat LM. Macular disease resembling adult foveomacular vitelliform dystrophy in older adults. Ophthalmology 1987; 94:362-6.
4. Feist RM, White MF Jr, Skalka H, Stone EM. Choroidal neovascularization in a patient with adult foveomacular dystrophy and a mutation in the retinal degeneration slow gene (Pro 210 Arg). Am J Ophthalmol 1994; 118:259-60.
5. Felbor U, Schilling H, Weber BH. Adult vitelliform macular dystrophy is frequently associated with mutations in the peripherin/RDS gene. Hum Mutat 1997; 10:301-9.
6. Gorin MB, Jackson KE, Ferrell RE, Sheffield VC, Jacobson SG, Gass JD, Mitchell E, Stone EM. A peripherin/retinal degeneration slow mutation (Pro-210-Arg) associated with macular and peripheral retinal degeneration. Ophthalmology 1995; 102:246-55.
7. Schatz P, Abrahamson M, Eksandh L, Ponjavic V, Andreasson S. Macular appearance by means of OCT and electrophysiology in members of two families with different mutations in RDS (the peripherin/RDS gene). Acta Ophthalmol Scand 2003; 81:500-7.
8. Wells J, Wroblewski J, Keen J, Inglehearn C, Jubb C, Eckstein A, Jay M, Arden G, Bhattacharya S, Fitzke F, Bird A. Mutations in the human retinal degeneration slow (RDS) gene can cause either retinitis pigmentosa or macular dystrophy. Nat Genet 1993; 3:213-8.
9. White K, Marquardt A, Weber BH. VMD2 mutations in vitelliform macular dystrophy (Best disease) and other maculopathies. Hum Mutat 2000; 15:301-8.
10. Allikmets R, Seddon JM, Bernstein PS, Hutchinson A, Atkinson A, Sharma S, Gerrard B, Li W, Metzker ML, Wadelius C, Caskey CT, Dean M, Petrukhin K. Evaluation of the Best disease gene in patients with age-related macular degeneration and other maculopathies. Hum Genet 1999; 104:449-53.
11. Kramer F, White K, Pauleikhoff D, Gehrig A, Passmore L, Rivera A, Rudolph G, Kellner U, Andrassi M, Lorenz B, Rohrschneider K, Blankenagel A, Jurklies B, Schilling H, Schutt F, Holz FG, Weber BH. Mutations in the VMD2 gene are associated with juvenile-onset vitelliform macular dystrophy (Best disease) and adult vitelliform macular dystrophy but not age-related macular degeneration. Eur J Hum Genet 2000; 8:286-92.
12. Seddon JM, Afshari MA, Sharma S, Bernstein PS, Chong S, Hutchinson A, Petrukhin K, Allikmets R. Assessment of mutations in the Best macular dystrophy (VMD2) gene in patients with adult-onset foveomacular vitelliform dystrophy, age-related maculopathy, and bull's-eye maculopathy. Ophthalmology 2001; 108:2060-7.
13. Edwards AO, Donoso LA, Ritter R 3rd. A novel gene for autosomal dominant Stargardt-like macular dystrophy with homology to the SUR4 protein family. Invest Ophthalmol Vis Sci 2001; 42:2652-63.
14. van Lith-Verhoeven JJ, Hoyng CB, van den Helm B, Deutman AF, Brink HM, Kemperman MH, de Jong WH, Kremer H, Cremers FP. The benign concentric annular macular dystrophy locus maps to 6p12.3-q16. Invest Ophthalmol Vis Sci 2004; 45:30-5.
15. Allikmets R, Singh N, Sun H, Shroyer NF, Hutchinson A, Chidambaram A, Gerrard B, Baird L, Stauffer D, Peiffer A, Rattner A, Smallwood P, Li Y, Anderson KL, Lewis RA, Nathans J, Leppert M, Dean M, Lupski JR. A photoreceptor cell-specific ATP-binding transporter gene (ABCR) is mutated in recessive Stargardt macular dystrophy. Nat Genet 1997; 15:236-46. Erratum in: Nat Genet 1997; 17:122.
16. Petrukhin K, Koisti MJ, Bakall B, Li W, Xie G, Marknell T, Sandgren O, Forsman K, Holmgren G, Andreasson S, Vujic M, Bergen AA, McGarty-Dugan V, Figueroa D, Austin CP, Metzker ML, Caskey CT, Wadelius C. Identification of the gene responsible for Best macular dystrophy. Nat Genet 1998; 19:241-7.
17. Sauer CG, Gehrig A, Warneke-Wittstock R, Marquardt A, Ewing CC, Gibson A, Lorenz B, Jurklies B, Weber BH. Positional cloning of the gene associated with X-linked juvenile retinoschisis. Nat Genet 1997; 17:164-70.
18. Stone EM, Lotery AJ, Munier FL, Heon E, Piguet B, Guymer RH, Vandenburgh K, Cousin P, Nishimura D, Swiderski RE, Silvestri G, Mackey DA, Hageman GS, Bird AC, Sheffield VC, Schorderet DF. A single EFEMP1 mutation associated with both Malattia Leventinese and Doyne honeycomb retinal dystrophy. Nat Genet 1999; 22:199-202.
19. Weber BH, Vogt G, Pruett RC, Stohr H, Felbor U. Mutations in the tissue inhibitor of metalloproteinases-3 (TIMP3) in patients with Sorsby's fundus dystrophy. Nat Genet 1994; 8:352-6.
20. Wrigley JD, Ahmed T, Nevett CL, Findlay JB. Peripherin/rds influences membrane vesicle morphology. Implications for retinopathies. J Biol Chem 2000; 275:13191-4.
21. Loewen CJ, Moritz OL, Molday RS. Molecular characterization of peripherin-2 and rom-1 mutants responsible for digenic retinitis pigmentosa. J Biol Chem 2001; 276:22388-96.
22. Goldberg AF, Moritz OL, Molday RS. Heterologous expression of photoreceptor peripherin/rds and Rom-1 in COS-1 cells: assembly, interactions, and localization of multisubunit complexes. Biochemistry 1995; 34:14213-9.
23. Kemp CM, Jacobson SG, Cideciyan AV, Kimura AE, Sheffield VC, Stone EM. RDS gene mutations causing retinitis pigmentosa or macular degeneration lead to the same abnormality in photoreceptor function. Invest Ophthalmol Vis Sci 1994; 35:3154-62.
24. Arnold JJ, Sarks JP, Killingsworth MC, Kettle EK, Sarks SH. Adult vitelliform macular degeneration: a clinicopathological study. Eye 2003; 17:717-26.
25. Jordan SA, Farrar GJ, Kenna P, Humphries P. Polymorphic variation within "conserved" sequences at the 3' end of the human RDS gene which results in amino acid substitutions. Hum Mutat 1992; 1:240-7.