Aberrant Splicing in the Ocular Albinism Type 1 Gene (OA1/GPR143) Is Corrected In Vitro by Morpholino Antisense Oligonucleotides

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An intronic point mutation was identified in theocular albinism type 1(OA1) gene (HUGO symbol,GPR143) in a family with the X-linked form of ocular albinism. Interestingly, the mutation creates a new acceptor splice site in intron 7 of theOA1gene. In addition to low levels of normally spliced mRNA product of theOA1gene, the patient samples contained also an aberrantly spliced mRNA with a 165 bp fragment of intron 7 (from position +750 to +914) inserted between exons 7 and 8. The abnormal transcript contained a premature stop codon and was unstable, as revealed by Northern blot analysis. We defined that mutation NC_000023.8:g.25288G>A generated a consensus binding motif for the splicing factor enhancer ASF/SF2, which most likely favored transcription of the aberrant mRNA. Furthermore, it activated a cryptic donor-splice site causing the inclusion between exons 7 and 8 of the 165 bp intronic fragment. Thus, the aberrant splicing is most likely explained by the generation of a de novo splicing enhancer motif. Finally, to rescue OA1 expression in the patient's melanocytes, we designed an antisense morpholino modified oligonucleotide complementary to the mutant sequence. The morpholino oligonucleotide (MO) was able to rescueOA1expression and restore the OA1 protein level in the patient's melanocytes through skipping of the aberrant inclusion. The use of MO demonstrated that the lack of OA1 was caused by the generation of a new splice site. Furthermore, this technique will lead to new approaches to correct splice site mutations that cause human diseases. Hum Mutat 27(5), 420-426, 2006. Published 2006 Wiley-Liss, Inc.†

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