Molecular dissection of Topologically Associating Domains (TADs)
Final Report Abstract
Mammalian genomes are organized into structural units called Topologically Associating Domains (TADs) that ensure adequate enhancer-promoter communication and gene expression. This critical function is mediated by a class of insulating regions named TAD boundaries that prevent aberrant regulatory interactions that might lead to ectopic gene activation. Accordingly, the disruption of TAD boundaries can lead to developmental disease or cancer. But despite the proven functional relevance of these regions, the molecular principles that operate at TAD boundaries had remained elusive. In this project, we perform analyses on 3D chromatin interactions and gene transcription in several mouse mutants to unravel the regulatory logic of TAD boundaries. We specifically focus on the boundaries at the Epha4 TAD, which are prototypical boundary regions associated with CTCF binding and which disruption results in ectopic gene expression and limb phenotypes. By analyzing transgenic mice carrying deletions of either the upstream or downstream Epha4 TAD boundaries, we demonstrate that the internal structure of a TAD can also contribute to its functional isolation. Individual deletions of CTCF binding sites (CBS) at boundaries revealed that these sites have distinct hierarchies in contributing to boundary insulation function. Furthermore, the combined deletion of CBS revealed that these sites cooperate redundantly and that divergent-oriented CBS, a recurrent signature of TAD boundaries, is not strictly required for robust insulation. Genome-wide analysis and in vivo validations also demonstrated the existence of chromatin loops mediated by non-convergently oriented CBS, formed by a process that we termed "loop interference”. By inverting and relocating TAD boundaries, we also show that boundary function can be largely independent of the surrounding genomic context. Finally, by correlating boundary strength, gene misexpression and phenotype penetrance, we demonstrate that TAD boundaries are quantitative modulators of gene expression and phenotypes. Overall, we reveal fundamental properties of TAD boundaries in orchestrating developmental gene expression and processes.
Publications
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Order and disorder: abnormal 3D chromatin organization in human disease. Briefings in Functional Genomics, 19(2), 128-138.
Anania, Chiara & Lupiáñez, Darío G.
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C/EBPβ regulates lipid metabolism and Pparg isoform 2 expression in alveolar macrophages. Science Immunology, 7(75).
Dörr; Dorothea; Obermayer; Benedikt; Weiner; January, Mikolaj; Zimmermann; Karin; Anania; Chiara; Wagner; Lisa, Katharina; Lyras; Ekaterini, Maria; Sapozhnikova; Valeriia; Lara-Astiaso; David; Prósper; Felipe; Lang; Roland; Lupiáñez; Darío, G.; Beule; Dieter; Höpken; Uta, E.; Leutz; Achim; Mildner & Alexander
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In vivo dissection of a clustered-CTCF domain boundary reveals developmental principles of regulatory insulation. Nature Genetics, 54(7), 1026-1036.
Anania, Chiara; Acemel, Rafael D.; Jedamzick, Johanna; Bolondi, Adriano; Cova, Giulia; Brieske, Norbert; Kühn, Ralf; Wittler, Lars; Real, Francisca M. & Lupiáñez, Darío G.
