Mapping of 3D genome topologies in human retina, RPE, PPCs and ROs
Non-coding structural variants (SVs) and regulatory single-nucleotide variants (SNVs) are still underrepresented in the mutation spectrum of IRD, often due to an interpretation gap. A missing link is the 3D interaction between cis-regulatory elements (CREs) and their target genes within topologically associating domains or TADs. Using chromatin interaction mapping (Hi-C) we have recently shown a differential 3D genome architecture of human retina and retinal pigment epithelium (RPE). DC5 will investigate if C-technologies on clinically accessible tissues (LCLs/fibroblasts) and on retinal stem cells (photoreceptor precursor cells/PPCs, retinal organoids/ROs) can be used to evaluate the effect of SVs causing IRD on the 3D genome. Following up on our previous Hi-C studies on retina, RPE, LCLs and fibroblasts, DC5 will perform an adapted Hi-C protocol (low-C) on PPCs and ROs. Apart from short-read sequencing-based Hi-C, DC5 will apply long-read sequencing-based Pore-C to retina, RPE, PPCs and ROs to generate a reference dataset that allows to unravel regulatory mechanisms within IRD loci, and to resolve complex SV. DC5 will assess the conservation of the 3D architecture between these cell types on a genomewide scale with special attention to adIRD loci. Next, DC5 will use the generated 3D data and perform C-technologies on available patient-derived cells as a phenotyping tool to map and interpret non-coding SVs found in genome data of unsolved IRD patients. Finally, C-technologies and insights from non-coding SV interpretation will ultimately improve genetic diagnoses in IRD.