Abstract
SummaryThe cause of autosomal-dominant retinitis pigmentosa (adRP), which leads to loss of vision and blindness, was investigated in families lacking a molecular diagnosis. A refined locus for adRP on Chr17q22 (RP17) was delineated through genotyping and genome sequencing, leading to the identification of structural variants (SVs) that segregate with disease. Eight different complex SVs were characterized in 22 adRP-affected families with >300 affected individuals. All RP17 SVs had breakpoints within a genomic region spanning YPEL2 to LINC01476. To investigate the mechanism of disease, we reprogrammed fibroblasts from affected individuals and controls into induced pluripotent stem cells (iPSCs) and differentiated them into photoreceptor precursor cells (PPCs) or retinal organoids (ROs). Hi-C was performed on ROs, and differential expression of regional genes and a retinal enhancer RNA at this locus was assessed by qPCR. The epigenetic landscape of the region, and Hi-C RO data, showed that YPEL2 sits within its own topologically associating domain (TAD), rich in enhancers with binding sites for retinal transcription factors. The Hi-C map of RP17 ROs revealed creation of a neo-TAD with ectopic contacts between GDPD1 and retinal enhancers, and modeling of all RP17 SVs was consistent with neo-TADs leading to ectopic retinal-specific enhancer-GDPD1 accessibility. qPCR confirmed increased expression of GDPD1 and increased expression of the retinal enhancer that enters the neo-TAD. Altered TAD structure resulting in increased retinal expression of GDPD1 is the likely convergent mechanism of disease, consistent with a dominant gain of function. Our study highlights the importance of SVs as a genomic mechanism in unsolved Mendelian diseases.
Highlights
Despite recent advances in next-generation sequencing, approximately 30%–40% of individuals with inherited retinal diseases (IRDs) lack a molecular diagnosis
To explore a convergent mechanism of disease, we investigated the effect of refined locus for adRP on Chr17q22 (RP17) structural variants (SVs) on three-dimensional (3D) chromatin organization that results in the compartmentalization of the genome into topologically associating domains (TADs) and the epigenetic landscape of the region
We demonstrate that altered TAD structure at the RP17 locus leads to ectopic retinal enhancer-gene interactions, consistent with a dominant gain of function
Summary
Despite recent advances in next-generation sequencing, approximately 30%–40% of individuals with inherited retinal diseases (IRDs) lack a molecular diagnosis. This is probably due to a combination of rare novel disease genes, which require large cohorts for validation, and previously intractable mutation classes, such as intronic variants, structural variants (SVs), and variants in regulatory regions.[1,2]. Autosomal-dominant RP (adRP) accounts for 25%–40% of cases, depending on the population studied, and has been associated with mutations in 30 genes, including CA4 (MIM: 114760) on Chr17q23.1 (RP17 [MIM: 600852])(see ‘‘RetNet’’ in Web Resources).[4,5] Following initial publications defining this locus[6,7] a variant in CA4 was implicated as the cause of adRP in families of South African origin, pathogenicity of the reported variant has been questioned because it has a population frequency of 4% in healthy controls in northern Sweden.[8,9,10] Subsequently reported CA4 variants in individuals with RP were identified by targeted Sanger sequencing and do not fully exclude variants in other genes as a cause of disease (Table S1)
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