Deep intronic mutation in OFD1, identified by targeted genomic next-generation sequencing, causes a severe form of X-linked retinitis pigmentosa (RP23)

Abstract

X-linked retinitis pigmentosa (XLRP) is genetically heterogeneous with two causative genes identified, RPGR and RP2. We previously mapped a locus for a severe form of XLRP, RP23, to a 10.71 Mb interval on Xp22.31-22.13 containing 62 genes. Candidate gene screening failed to identify a causative mutation, so we adopted targeted genomic next-generation sequencing of the disease interval to determine the molecular cause of RP23. No coding variants or variants within or near splice sites were identified. In contrast, a variant deep within intron 9 of OFD1 increased the splice site prediction score 4 bp upstream of the variant. Mutations in OFD1 cause the syndromic ciliopathies orofaciodigital syndrome-1, which is male lethal, Simpson-Golabi-Behmel syndrome type 2 and Joubert syndrome. We tested the effect of the IVS9+706A>G variant on OFD1 splicing in vivo. In RP23 patient-derived RNA, we detected an OFD1 transcript with the insertion of a cryptic exon spliced between exons 9 and 10 causing a frameshift, p.N313fs.X330. Correctly spliced OFD1 was also detected in patient-derived RNA, although at reduced levels (39%), hence the mutation is not male lethal. Our data suggest that photoreceptors are uniquely susceptible to reduced expression of OFD1 and that an alternative disease mechanism can cause XLRP. This disease mechanism of reduced expression for a syndromic ciliopathy gene causing isolated retinal degeneration is reminiscent of CEP290 intronic mutations that cause Leber congenital amaurosis, and we speculate that reduced dosage of correctly spliced ciliopathy genes may be a common disease mechanism in retinal degenerations.

The role of primary cilia in the development and disease of the retina

“…36 Mutations in OFD1 cause oro-facial-digital syndrome, non-syndromic RP, and RP as a feature of Joubert syndrome (JBTS). [37][38][39] ofd1 morphant zebrafish occasionally develop retinal coloboma.…”

Improving molecular diagnosis in epilepsy by a dedicated high-throughput sequencing platform

“…This point was taken with extreme caution as it could not be excluded, a priori, that a novel mutation might cause a totally different phenotype with respect to the ones known to be associated with the same gene. 13 Some of the remaining alterations, even if predicted damaging by at least one tool, have been discarded when they did not segregate with the epileptic phenotype in familial cases (ie, MAGI2 in case 5-A that was inherited by the healthy mother, whereas KCNQ2 was considered causative because it was inherited by the affected father).…”

Generation of Hypertension-Associated STK39 Polymorphism Knockin Cell Lines With the Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 System

“…27 We first investigated whether alternative splicing of STK39 occurred in knockin cells. Amplicon sizes of reverse transcription PCR spanning exons 5 to 7 were similar between WT and knockin cell lines (Figure 2A).…”