Modified U1 snRNA and antisense oligonucleotides rescue splice mutations in SLC26A4 that cause hereditary hearing loss.

One of most important factors for messenger RNA (mRNA) transcription is the spliceosomal component U1 small nuclear RNA (snRNA), which recognizes 5' splicing donor sites at specific regions in pre-mRNA. Mutations in these sites disrupt U1 snRNA binding and cause abnormal splicing. In this study, we investigated mutations at splice sites in SLC26A4 (HGNC 8818), one of the major causative genes of hearing loss, which may result in the synthesis of abnormal pendrin, the channel protein encoded by the gene. Seventeen SLC26A4 variants with mutations in the U1 snRNA binding sites were assessed by minigene splicing assays, and 11 were found to result in abnormal splicing. Interestingly, eight of the 11 pathogenic mutations were intronic, suggesting the importance of conserved sequences at the intronic splice site. The application of modified U1 snRNA effectively rescued the abnormal splicing for most of these mutations. Although three were cryptic mutations, they were rescued by cotransfection of modified U1 snRNA and modified antisense oligonucleotides. Our results demonstrate the important role of snRNA in SLC26A4 mutations, suggesting the therapeutic potential of modified U1 snRNA and antisense oligonucleotides for neutralizing the pathogenic effect of the splice-site mutations that may result in hearing loss.