Lupiáñez Darío G, Kraft Katerina, Heinrich Verena, Krawitz Peter, Brancati Francesco, Klopocki Eva, Horn Denise, Kayserili Hülya, Opitz John M, Laxova Renata, Santos-Simarro Fernando, Gilbert-Dussardier Brigitte, Wittler Lars, Borschiwer Marina, Haas Stefan A, Osterwalder Marco, Franke Martin, Timmermann Bernd, Hecht Jochen, Spielmann Malte, Visel Axel, Mundlos Stefan
Max Planck Institute for Molecular Genetics, RG Development & Disease, 14195 Berlin, Germany; Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, 13353 Berlin, Germany.
Cell. 2015 May 21;161(5):1012-1025. doi: 10.1016/j.cell.2015.04.004. Epub 2015 May 7.
Mammalian genomes are organized into megabase-scale topologically associated domains (TADs). We demonstrate that disruption of TADs can rewire long-range regulatory architecture and result in pathogenic phenotypes. We show that distinct human limb malformations are caused by deletions, inversions, or duplications altering the structure of the TAD-spanning WNT6/IHH/EPHA4/PAX3 locus. Using CRISPR/Cas genome editing, we generated mice with corresponding rearrangements. Both in mouse limb tissue and patient-derived fibroblasts, disease-relevant structural changes cause ectopic interactions between promoters and non-coding DNA, and a cluster of limb enhancers normally associated with Epha4 is misplaced relative to TAD boundaries and drives ectopic limb expression of another gene in the locus. This rewiring occurred only if the variant disrupted a CTCF-associated boundary domain. Our results demonstrate the functional importance of TADs for orchestrating gene expression via genome architecture and indicate criteria for predicting the pathogenicity of human structural variants, particularly in non-coding regions of the human genome.
哺乳动物基因组被组织成兆碱基规模的拓扑相关结构域（TADs）。我们证明，TADs的破坏可重新连接远程调控架构并导致致病表型。我们表明，不同的人类肢体畸形是由缺失、倒位或重复导致跨越TAD的WNT6/IHH/EPHA4/PAX3基因座结构改变引起的。使用CRISPR/Cas基因组编辑技术，我们构建了具有相应重排的小鼠。在小鼠肢体组织和患者来源的成纤维细胞中，与疾病相关的结构变化均导致启动子与非编码DNA之间的异位相互作用，并且通常与Epha4相关的一组肢体增强子相对于TAD边界位置错误，并驱动该基因座中另一个基因的异位肢体表达。只有当变异破坏了与CTCF相关的边界域时，这种重新连接才会发生。我们的结果证明了TADs通过基因组架构协调基因表达的功能重要性，并指出了预测人类结构变异致病性的标准，特别是在人类基因组的非编码区域。
