3D Genome Architecture in Congenital Disease

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One of the major scientific breakthoughs in genomics of the last decade has been the discovery that the genome folds in the threedimensional space in the nucleus and that this is directly related to gene regulation. In this proposal I propose to take disease oriented genome analysis to another level by combining the study 3D genome architecture with comprehensive genome analysis, thereby addressing one of the major problems in our current understanding of rare diseases. I hypothesize that many so far undiagnosed cases are in fact caused by abnormal gene regulation due to mutations that alter genomic architecture thereby changing regulatory enhancer-promoter interactions. I propose to detect and investigate such novel mutational mechanisms genome wide using a chromosome conformation capture (HiC) in cells from patients with congenital limb malformations. HiC will be used to identify aberrant enhancerpromoter contacts that might influence developmental gene expression. These data will be compared to whole genome analysis with short (Illumina) and long (PacBio) read technology to identify corresponding changes including variants of sequence and structure as well as repetitive sequences that might change 3D genome architecture. Changes with potential pathogenicity will be investigated in vivo in mice using an adapted Crispr/Cas9 protocol or in vitro in fibroblasts or iPSCs generated from these cells.