The human genome is segmented into topologically associating domains (TADs), but the role of this conserved organization during transient changes in gene expression is not known. Here we describe the distribution of progestin-induced chromatin modifications and changes in transcriptional activity over TADs in T47D breast cancer cells. Using ChIP-seq (chromatin immunoprecipitation combined with high-throughput sequencing), Hi-C (chromosome capture followed by high-throughput sequencing), and three-dimensional (3D) modeling techniques, we found that the borders of the ∼2000 TADs in these cells are largely maintained after hormone treatment and that up to 20% of the TADs could be considered as discrete regulatory units where the majority of the genes are either transcriptionally activated or repressed in a coordinated fashion. The epigenetic signatures of the TADs are homogeneously modified by hormones in correlation with the transcriptional changes. Hormone-induced changes in gene activity and chromatin remodeling are accompanied by differential structural changes for activated and repressed TADs, as reflected by specific and opposite changes in the strength of intra-TAD interactions within responsive TADs. Indeed, 3D modeling of the Hi-C data suggested that the structure of TADs was modified upon treatment. The differential responses of TADs to progestins and estrogens suggest that TADs could function as “regulons” to enable spatially proximal genes to be coordinately transcribed in response to hormones.

How genes are regulated in the context of chromatin is a central question of biology. Steroid hormones control gene expression via interaction of their receptors with target sequences on DNA but can also activate cytoplasmic signaling cascades. Here we report that rapid Erk activation by progestins participates in induction of target genes by preparing the chromatin for transcription. Five minutes after hormone treatment, Erk activation leads to phosphorylation of the progesterone receptor (PR), activation of Msk1, and recruitment of a complex of the three proteins to a nucleosome on the MMTV promoter. Msk1 phosphorylates histone H3, leading to displacement of HP1gamma and recruitment of Brg1 and RNA polymerase II. Cell-free experiments show a direct interaction between PR, Erk, and Msk1 and support the importance of H3 phosphorylation for nucleosome remodeling. Inhibition of Msk1 activation blocks recruitment of the kinase complex, H3 phosphorylation, and HP1gamma displacement, thus precluding remodeling and induction of the promoter.

Polycomb-group proteins are transcriptional repressors with essential roles in embryonic development. Polycomb repressive complex 2 (PRC2) contains the methyltransferase activity for Lys27. However, the role of other histone modifications in regulating PRC2 activity is just beginning to be understood. Here we show that direct recognition of methylated histone H3 Lys36 (H3K36me), a mark associated with activation, by the PRC2 subunit Phf19 is required for the full enzymatic activity of the PRC2 complex. Using NMR spectroscopy, we provide structural evidence for this interaction. Furthermore, we show that Phf19 binds to a subset of PRC2 targets in mouse embryonic stem cells and that this is required for their repression and for H3K27me3 deposition. These findings show that the interaction of Phf19 with H3K36me2 and H3K36me3 is essential for PRC2 complex activity and for proper regulation of gene repression in embryonic stem cells.

SummaryThe unicellular ancestor of animals had a complex repertoire of genes linked to multicellular processes. This suggests that changes in the regulatory genome, rather than in gene innovation, were key to the origin of animals. Here, we carry out multiple functional genomic assays in Capsaspora owczarzaki, the unicellular relative of animals with the largest known gene repertoire for transcriptional regulation. We show that changing chromatin states, differential lincRNA expression, and dynamic cis-regulatory sites are associated with life cycle transitions in Capsaspora. Moreover, we demonstrate conservation of animal developmental transcription-factor networks and extensive network interconnection in this premetazoan organism. In contrast, however, Capsaspora lacks animal promoter types, and its regulatory sites are small, proximal, and lack signatures of animal enhancers. Overall, our results indicate that the emergence of animal multicellularity was linked to a major shift in genome cis-regulatory complexity, most notably the appearance of distal enhancer regulation.