Molecular dissection of Topologically Associating Domains (TADs)

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3D folding is an inherent property of the vertebrate genome in order to accommodate the roughly 2m of DNA that a single cell contains in such a tiny space as the nucleus. This folding, far from being a random process, appears to be highly predetermined and strongly influences gene expression. At the subchromosomal scale, our genome is partitioned in self-interacting units called Topologically Associating Domains (TADs), ranging from a few hundreds of base pairs to several megabases. TADs are delimited by so-called boundary regions that isolate them from the rest of the genome, thus promoting contacts between the loci contained within them. It has been proposed that TADs represent a fundamental structural unit of the genome, which is thought to guide regulatory elements to their cognate genes. Recently, the disruption of TADs and their boundaries was shown to be associated to human disorders such as congenital malformations or cancer. Despite the biological relevance of TADs, the essential components required to build such a fundamental structure remain elusive. In this proposal, I aim to molecularly dissect TAD domains and their boundary regions, in order to understand how these structures can be formed and influenced by their genomic context. By combining cutting-edge technologies such as Capture-HiC or CRISPR/Cas to generate mice carrying specific mutations, I am to: 1) evaluate the contribution of TAD boundaries in domain organization; 2) determine the minimal region to confer boundary function and 3) evaluate the impact of genomic context on TAD boundary function. The potential results derived from this proposal would advance in our understanding of the general principles of chromatin organization and how their alteration can originate abnormal phenotypes.