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Francesco Paolo Ruberto, Sara Balzano, Adva Kimchi, Prasanthi Namburi, Tamar Ben-Yosef, Eyal Banin, Dror Sharon, Carlo Rivolta; A heterozygous deletion of a noncoding part of the PRPF31 gene causes retinitis pigmentosa via reduced gene expression. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5421. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Pathogenic variants in the gene encoding the splicing factor PRPF31 can cause autosomal dominant retinitis pigmentosa (adRP) with incomplete penetrance. Mutations are usually frameshift of nonsense changes that lead to the reduction of the overall level of encoded protein, a phenomenon that can be reversed in unaffected carriers by overexpression of the wild-type allele. This study aims at validating this haploinsufficient mechanism of pathogenesis through the characterization of a deletion that encompasses a regulatory region of PRPF31, but leaves its coding sequence completely intact.
We studied a family of Ashkenazi Jewish descent with a history of non-penetrant adRP, using whole exome sequencing (WES), Sanger sequencing, and multiplex ligation-dependent probe amplification (MLPA) of the PRPF31 gene. Five members of the family, including an unaffected carrier, underwent a complete ophthalmic evaluation and peripheral blood drawing, for extraction of genomic DNA. Lymphocytes from participants as well as from control subjects were immortalized and used to assess mRNA expression by q-PCR.
WES analysis of the three affected individuals and the unaffected carrier did not reveal any potential pathogenic mutations, but linkage analysis was compatible with PRPF31 as the cause of disease. MLPA analysis of the PRPF31 gene revealed a putative deletion in exon 1, which is noncoding, in two affected siblings. At the genomic level, the mutation was characterized by serial very-short-range PCRs, followed by a long-range PCR, and finally by a diagnostic PCR encompassing the deletion area and providing a binary output (presence/absence of PCR product). This allowed to characterize the mutation as a 3.5-kilobase deletion encompassing PRPF31’s core promoters and its first exon. The deletion affected as well exon 1 and 2 of TFPT, a gene transcribed in the opposite direction with respect to PRPF31. At the RNA level, transcript analysis showed that patients expressed lower levels of PRPF31 mRNA, compared to controls. The unaffected carrier of the deletion had intermediate levels of mRNA.
Our study shows that PRPF31-associated adRP can be caused by mutations in noncoding parts of the gene, leading in turn to reduced mRNA expression and to disease, via a haploinsufficient mechanism.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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