Determining the functional relationship between epigenetic and physical chromatin domains in Drosophila

Eukaryotic genomes are hierarchically organized into topologically associating domains (TADs) that regulate genome function. In Drosophila , TADs strongly correlate with epigenetic domains marked by either active or repressive chromatin marks. This tight correlation may suggest causality between the deposition of the epigenomic marks and formation of TADs. However, it is still unknown whether the epigenome is a major driving force for TAD formation and structure and what is their functional importance. Here we directly address these questions by perturbing the position of Polycomb response elements (PREs), which act as nucleation sites for Polycomb group proteins (PcG) to deposit H3K27me3 and form epigenetic domains in the dac TAD. First, we remove the two PREs in the dac domain wiping out the H3K27me3 mark from the TAD and study the consequences on the physical property of the domain. Second, by shifting the position of one of the PREs, we create an ectopic PcG epigenetic domain and analyze how this affects TAD formation. Together, these two experimental perturbations indicate that, although TADs correlate very well with chromatin states in Drosophila , the epigenomic signature of Polycomb TADs does not play a causal role in the establishment of TADs. On the other hand, physical domains have an important impact on the formation of epigenetic domains, as they can restrict spreading of the repressive histone marks and of PRE loops.