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Evidence for Pathogenicity of Atypical Splice Mutations in Autosomal Dominant Polycystic Kidney Disease

Kiarong Wang^*, Xiao Zhao^*,, Shelly Chan^*, Onur Cil^*, Ning He^*, Xuewen Song^*, Andrew D. Paterson, and York Pei^*

^* Divisions of Nephrology and Genomic Medicine, University Health Network and University of Toronto, Toronto, Ontario Canada; Schulich School of Medicine, University of Western Ontario, London, Ontario, Canada; Program in Genomic Biology, Hospital for Sick Children, Toronto, Ontario, Canada

Correspondence: Dr. York Pei, Division of Nephrology, University Health Network, 8N838, 585 University Avenue, Toronto, Ontario, Canada M5G 2N2. Phone: 416-340-4257; Fax: 416-340-4999; E-mail: york.pei{at}uhn.on.ca

Background and objectives: Mutation-based molecular diagnostics of autosomal dominant polycystic kidney disease (ADPKD) is complicated by locus and allelic heterogeneity, large multi-exon gene structure and duplication in PKD1, and a high level of unclassified variants. Comprehensive screening of PKD1 and PKD2 by two recent studies have shown that atypical splice mutations account for 3.5% to 5% of ADPKD. We evaluated the role of bioinformatic prediction of atypical splice mutations and determined the pathogenicity of an atypical PKD2 splice variant from a multiplex ADPKD (TOR101) family.

Design, setting, participants, & measurements: Using PubMed, we identified 17 atypical PKD1 and PKD2 splice mutations. We found that bioinformatics analysis was often useful for evaluating the pathogenicity of these mutations, although RT-PCR is needed to provide the definitive proof.

Results: Sequencing of both PKD1 and PKD2 in an affected subject of TOR101 failed to identify a definite mutation, but revealed several UCVs, including an atypical PKD2 splice variant. Linkage analysis with microsatellite markers indicated that TOR101 was PKD2-linked and IVS8 + 5GA was shown to cosegregate only with affected subjects. RT-PCR of leukocyte mRNA from an affected subject using primers from exons 7 and 9 revealed six splice variants that resulted from activation of different combinations of donor and acceptor cryptic splice sites, all terminating with premature stop codons.

Conclusions: The data provide strong evidence that IVS8 + 5GA is a pathogenic mutation for PKD2. This case highlights the importance of functional analysis of UCVs.