title: Erosive vitreoretinopathy and wagner disease are caused by intronic mutations in CSPG2/Versican that result in an imbalance of splice variants.
abstract: OBJECTIVE
  Linkage intervals for erosive vitreoretinopathy (ERVR) and Wagner disease previously were found to overlap at 5q14.3. In a Japanese family with Wagner disease, a CSPG2/Versican splice site mutation (c.4004-2A-->G) was recently reported that resulted in a 39-nucleotide exon 8 in-frame deletion. We investigated whether CSPG2/Versican was mutated in six Dutch families and one Chinese family with Wagner disease and in a family with ERVR.
  
  
  METHODS
  In all families, extensive ophthalmic examinations, haplotype analysis of the 5q14.3 region, and sequence analysis of CSPG2/Versican were performed. The effects of splice site mutations were assessed by reverse transcription-polymerase chain reaction (RT-PCR) and real-time quantitative RT-PCR (QPCR).
  
  
  RESULTS
  Three novel intron 7 sequence variants (c.4004-5T-->C, c.4004-5T-->A, c.4004-1G-->A) were identified in seven families. The c.4004-5T-->C variant was identified in four families with Wagner disease and a family with ERVR. The families were shown to carry the same 5q14.3 haplotype, strongly suggesting that this is a common Dutch founder variant. All three changes segregated with the disease in the respective families and were absent in 250 healthy individuals. In patients with the c.4004-5T-->A and c.4004-1G-->A variants, RT-PCR analysis of CSPG2/Versican showed activation of a cryptic splice site resulting in a 39-nt exon 8 in-frame deletion in splice variant V0. QPCR revealed a highly significant (P < 0.0001) and consistent increase of the V2 (>38-fold) and V3 (>12-fold) splice variants in all patients with intron 7 nucleotide changes and in a Chinese Wagner disease family, in which the genetic defect remains to be found.
  
  
  CONCLUSIONS
  Wagner disease and ERVR are allelic disorders. Seven of the eight families exhibit a variant in intron 7 of CSPG2/Versican. The conspicuous clustering of sequence variants in the splice acceptor site of intron 7 and the consistent upregulation of the V2 and V3 isoforms strongly suggest that Wagner disease and ERVR may belong to a largely overlooked group of diseases that are caused by mRNA isoform balance shifts, representing a novel disease mechanism.
authors.affiliation: Department of Human Genetics, Radboud University Nijmegen Medical Centre, The Netherlands.
authors.forename: Arijit
authors.initials: A
authors.lastname: Mukhopadhyay
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Nikopoulos
authors.forename: Alessandra
authors.initials: A
authors.lastname: Maugeri
authors.forename: Arjan P M
authors.initials: AP
authors.lastname: de Brouwer
authors.forename: C Eric
authors.initials: CE
authors.lastname: van Nouhuys
authors.forename: Camiel J F
authors.initials: CJ
authors.lastname: Boon
authors.forename: Rahat
authors.initials: R
authors.lastname: Perveen
authors.forename: Hester A A
authors.initials: HA
authors.lastname: Zegers
authors.forename: Dienke
authors.initials: D
authors.lastname: Wittebol-Post
authors.forename: Pieter R
authors.initials: PR
authors.lastname: van den Biesen
authors.forename: Saskia D
authors.initials: SD
authors.lastname: van der Velde-Visser
authors.forename: Han G
authors.initials: HG
authors.lastname: Brunner
authors.forename: Graeme C M
authors.initials: GC
authors.lastname: Black
authors.forename: Carel B
authors.initials: CB
authors.lastname: Hoyng
authors.forename: Frans P M
authors.initials: FP
authors.lastname: Cremers
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D002648:Child; D002675:Child, Preschool; D011508:Chondroitin Sulfate Proteoglycans / Q000235:genetics*; D002895:Chromosomes, Human, Pair 5 / Q000235:genetics; D005128:Eye Diseases / Q000235:genetics*; D005260:Female; D008040:Genetic Linkage; D014644:Genetic Variation; D005838:Genotype; D006239:Haplotypes; D006801:Humans; D007438:Introns / Q000235:genetics*; D037181:Lectins, C-Type / Q000235:genetics*; D008297:Male; D009154:Mutation; D010375:Pedigree; D022821:RNA Splice Sites / Q000235:genetics*; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics; D012164:Retinal Diseases / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053675:Versicans; D014822:Vitreous Body
chemical_list: D011508:Chondroitin Sulfate Proteoglycans; D037181:Lectins, C-Type; D022821:RNA Splice Sites; D012333:RNA, Messenger; C508322:VCAN protein, human; D053675:Versicans