Novel VCAN mutations and evidence for unbalanced alternative splicing in the pathogenesis of Wagner syndrome

Abstract

Wagner syndrome (WS) is an autosomal dominant vitreoretinopathy affecting various ocular features and is caused by mutations in the canonical splice sites of the VCAN gene, which encodes the large chondroitin sulfate proteoglycan, versican. We report the identification of novel splice acceptor and donor-site mutations (c.4004 À1G4C and c.9265 þ 2T4A) in two large WS families from France and the United Kingdom. To characterize their pathogenic mechanisms we performed qRT-PCR experiments on RNA from patient-derived tissues (venous blood and skin fibroblasts). We also analyzed RNA from the original Swiss family reported by Wagner (who has the previously reported c.9265 þ 1G4A mutation). All three mutations resulted in a quantitative increase of transcript variants lacking exons 7 and/or 8. However, the magnitude of the increase varied between tissues and mutations. We discuss altered balance of VCAN splice variants in combination with reduction in glycosaminoglycan protein modifications as possible pathogenic mechanisms.

Understanding disease pleiotropy: From puzzle to solution

“…Altering the f netuning of the relative abundance of splicing isoforms could also be problematic. T e contribution of VCAN unbalanced alternative splicing in the pathogenesis of Wagner syndrome is consistent with this view (10). Interestingly, exons undergoing basal exon skipping are strong candidates for splice-switching oligonucleotide therapies, which are designed to bypass protein truncation.…”

The versican-hyaluronan complex provides an essential extracellular matrix niche for Flk1+ hematoendothelial progenitors

“…We found a stronger association of exon 8-containing Vcan transcripts than exon 7containing transcripts with blood islands at gastrulation and exon 7 transcripts were absent at E8.5. Autosomal dominant splice site mutations affecting exon 8 (leading to its exclusion) and exon 8 deletions in humans cause Wagner syndrome (Kloeckener-Gruissem et al, 2006;Kloeckener-Gruissem et al, 2013;Rothschild et al, 2013), which is characterized by impaired vision and defects of the ocular vitreous and retina, but lacks consistent extra-ocular manifestations. Exon 7 inactivation in mice led to specific neural anomalies and subtle cardiac anomalies (Burns et al, 2014;Dours-Zimmermann et al, 2009).…”

A potential mouse model for the erosive vitreoretinopathy of Wagner disease      

“…Whether the patho-mechanism is also comparable remains to be investigated because the mouse strain carries an expression null allele [16], while human patients still may express the two smallest splice variants V2 and V3 from the affected allele. It has previously been suggested that the splice defect might result in an imbalanced ratio of transcript isoforms that may lead to an imbalance of versican protein isoforms [9] [4] [11]. The observation that retinal morphology was not altered in hdf +/-mice suggests that the abnormal ERG response may be caused by cellular signalling defects rather than cell death.…”

“…Furthermore, versican has been found as a component of the vitreous [5] [6]. To date all identified mutations in patients with Wagner disease map to the canonical splice sites bordering exon 8 [7] [8] [9] [10] [11] [12] [13]. Under normal conditions, 4 alternative splice isoforms are produced, which differ in the presence of exons 7 and 8 (variant V0 contains exon 7 and 8, V1 lacks exon 7, V2 lacks exon 8 and V3 lacks exon 7 and 8).…”