ИСТИНА

In this study, we examine chromatin folding in Drosophila to uncover principles of Topologically Associating Domains (TADs) formation. To this end, we performed comparative Hi-C analysis in four cell lines of different origin and developmental stages (S2, Kc167, BG3 and OSC), annotated TADs genome-wide using the two-step Armatus-based approach and analyzed epigenetic features of TADs, TAD boundaries and inter-TAD regions. To precisely relate transcription profile to TAD positions genome-wide, we performed Poly(A)+ transcriptome sequencing of the studied cell lines. Together with the existing epigenetic tracks from modENCODE, our data allowed us to make several important observations and develop a model of the Drosophila chromatin packaging into TADs. We found that TAD boundaries and extended inter-TADs regions in all examined cell lines are strongly enriched with acetylated histones, highly transcribed housekeeping genes, active enhancers and correspond to decompacted interbands of polytene chromosomes. Contrary to previous reports, we observed an almost uniform distribution of insulator protein dCTCF between TADs and inter-TADs, and increased level of insulator protein Su(Hw) within TADs. Using a novel strategy for the analysis of the Hi-C data, we showed that active chromatin state and transcription, but not the presence of the aforementioned insulator proteins quantitatively interfere with chromatin fiber packaging into TADs. We found that active transcription and active chromatin marks can predict inter-TADs and TAD boundaries reasonably well; moreover, including the profiles of dCTCF and Su(Hw) does not improve this prediction, that strongly suggests against their central role in demarcating TAD boundaries. Finally, we propose, and verify by polymer simulations, a simple model of TAD self-assembly. In our model numerous transient interactions between non-acetylated nucleosomes of inactive chromatin lead to the stochastic formation of TADs. On the contrary, regions harboring actively transcribed (predominantly housekeeping) genes remain unfolded, likely because the inter-nucleosomal interactions are suppressed by a high level of histone acetylation.