Suzanne de Bruijn

181 Structural variants cause ectopic enhancer-gene contact in retinitis pigmentosa Inbothexperimentalmodels, the expression levels of GDPD1 were significantly increased compared to controls. YPEL2 was increased in NL-SV1 only ( Figure 5C ), whereas SMG8 was increased in UK-SV2 ( Figure 5E ), which correlates with our TAD modelling and Hi-C experimental data for UK-SV2 ROs ( Figure 5B , 5D , S9 ). To further explore the tissue specific effect of this transcriptional upregulation, we performed the same qPCR assays on fibroblasts of the same individuals. None of these genes had increased expression levels in affected individuals compared to controls (data not shown). DISCUSSION Previous genetic studies of adRP families mapping to the RP17-locus have implicated missense variants in the CA4 gene as the cause of disease, or have been unable to confirm pathogenicity. 9,10 Here, we describe the discovery of SVs as the cause of adRP at the RP17-locus in a large number of families, suggesting this is a previously unrecognized major locus for adRP. Our results show how complex rearrangements can result in the disruption of 3D genome architecture, the re-wiring of enhancer-promoter interactions and consequent gene misexpression. Following identification of SVs in NL1 and UK1 using short-read WGS, our search for similar complex SVs in the RP17 genomic interval of genetically unexplained adRP families identified six other complex SVs that segregated with disease. SVs are a major source of normal variation in the human genome and are often benign; 29,30 however, none of the RP17-SVs are found in the population database gnomAD 31 or the Database of Genomic Variants (DGV). 32 Although overlapping canonical SVs (deletions and duplications) have been identified, they do not have breakpoints within the YPEL2 - LINC01476 region, as observed for all RP17-SVs reported in this study. This is in line with observations that different SVs can have different consequences, depending on the characteristics of specific SVs in local 3D chromatin and epigenetic context. 33,34 Base level resolution of breakpoint junctions and interrogation of the DNA sequence signatures revealed the mechanisms of the chromosomal rearrangements. Repetitive elements are key factors in facilitating unequal crossover of genomic segments or to providemicrohomologythat inducesforkstallingandsubsequenttemplateswitching. 35,36 Consistent with this model, repetitive elements were present in the flanking sequences of breakpoint junctions. In addition, microhomology, larger stretches of homology and small insertions-deletions were found at all breakpoints. 36-38 Therefore, repetitive elements may explain why the RP17-locus is prone to such structural variation, which is