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A. Mukhopadhyay, K. Nikopoulos, A. Maugeri, E. van Nouhuys, A.P. de Brouwer, D. Wittebol–Post, G.C. M. Black, C.B. Hoyng, F.P. M. Cremers; CSPG2/Versican Intron 7 Mutations Cause Wagner Disease Due to an Imbalance Between Splice Variants . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4635.
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In a single Japanese family with Wagner disease, an autosomal dominant vitreoretinopathy, a CSPG2/Versican splice site mutation (c.4004–2A>G) was reported which resulted in an apparently mild in–frame 39–nt deletion of exon 8 (Miyamoto et. al. IOVS 46:2726–2735, 2005). The purpose of this study was to analyse the nucleotide sequence of the CSPG2/Versican gene and its mRNA expression in 6 large Dutch Wagner disease families.
Semi–automated sequence analysis of CSPG2/Versican exons 2–15, including 50 bp of flanking intronic sequences. Quantitative RT–PCR of mRNA was performed of the splice variants of CSPG2/Versican using the MyiQ single color Real–Time detection system (Biorad).
Three different sequence variants (c.4004–5T>C, c.4004–5T>A, c.4004–1G>A) were identified in intron 7. The c.4004–5T>C variant was identified in 4 Dutch families which were shown to carry the same 5q14.3 haplotype, strongly suggesting that this represents a common Dutch founder variant. Regular RT–PCR analysis showed the variable activation of a cryptic splice site resulting in an in–frame 39–nt deletion of exon 8 in splice variants V0 and V1. Quantitative RT–PCR revealed a consistent increase of the V2 (>40–fold) and V3 (>10–fold) splice variants, in all patients with intron 7 splice site sequence changes.
The conspicuous clustering of sequence variants in the splice acceptor site of intron 7 and the consistent upregulation of the V2 and V3 mRNA isoforms, lead us to believe that Wagner disease is caused by an imbalance of CSPG2/Versican protein isoforms whichcan only be triggered by intronic mutations.
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