CTCF, BEAF-32 and CP190 are not required for the initial establishment of TADs in early Drosophila embryos, but have locus specific roles

Abstract

The boundaries of Topologically-Associating Domains (TADs) are delimited by insulators and active promoters, however how they are initially established during embryogenesis remains unclear. Here, we examined this during the first hours of Drosophila embryogenesis. DNA-FISH on individual embryos indicates that domains form during zygotic genome activation (ZGA), but have extensive cell-to-cell heterogeneity compared to later stages. Most newly formed boundaries are occupied by combinations of CTCF, BEAF- 32 and/or CP190. Depleting each insulator from chromatin revealed that TADs can still establish during ZGA, although with lower insulation, with particular boundaries being more sensitive. Some weakened boundaries have aberrant gene expression, however the majority of mis-expressed genes have no obvious relationship to changes in domain-boundary insulation. Deletion of an active promoter (thereby blocking transcription) at one boundary had a greater impact compared to deleting the insulator-bound region itself. These results suggest cross-talk between insulators and transcription might reinforce domain formation during embryogenesis.