SummaryMouse embryonic stem (ES) cells grown in serum exhibit greater heterogeneity in morphology and expression of pluripotency factors than ES cells cultured in defined medium with inhibitors of two kinases (Mek and GSK3), a condition known as “2i” postulated to establish a naive ground state. We show that the transcriptome and epigenome profiles of serum- and 2i-grown ES cells are distinct. 2i-treated cells exhibit lower expression of lineage-affiliated genes, reduced prevalence at promoters of the repressive histone modification H3K27me3, and fewer bivalent domains, which are thought to mark genes poised for either up- or downregulation. Nonetheless, serum- and 2i-grown ES cells have similar differentiation potential. Precocious transcription of developmental genes in 2i is restrained by RNA polymerase II promoter-proximal pausing. These findings suggest that transcriptional potentiation and a permissive chromatin context characterize the ground state and that exit from it may not require a metastable intermediate or multilineage priming.

NuRD (nucleosome remodeling and histone deacetylase) is a versatile multi-protein complex with roles in transcription regulation and the DNA damage response. Here, we show that ZMYND8 bridges NuRD to a number of putative DNA-binding zinc finger proteins. The MYND domain of ZMYND8 directly interacts with PPPLΦ motifs in the NuRD subunit GATAD2A. Both GATAD2A and GATAD2B exclusively form homodimers and define mutually exclusive NuRD subcomplexes. ZMYND8 and NuRD share a large number of genome-wide binding sites, mostly active promoters and enhancers. Depletion of ZMYND8 does not affect NuRD occupancy genome-wide and only slightly affects expression of NuRD/ZMYND8 target genes. In contrast, the MYND domain in ZMYND8 facilitates the rapid, poly(ADP-ribose)-dependent recruitment of GATAD2A/NuRD to sites of DNA damage to promote repair by homologous recombination. Thus, these results show that a specific substoichiometric interaction with a NuRD subunit paralogue provides unique functionality to distinct NuRD subcomplexes.

BackgroundTranscription factor-dependent cellular reprogramming is integral to normal development and is central to production of induced pluripotent stem cells. This process typically requires pioneer transcription factors (TFs) to induce de novo formation of enhancers at previously closed chromatin. Mechanistic information on this process is currently sparse.ResultsHere we explore the mechanistic basis by which GATA3 functions as a pioneer TF in a cellular reprogramming event relevant to breast cancer, the mesenchymal to epithelial transition (MET). In some instances, GATA3 binds previously inaccessible chromatin, characterized by stable, positioned nucleosomes where it induces nucleosome eviction, alters local histone modifications, and remodels local chromatin architecture. At other loci, GATA3 binding induces nucleosome sliding without concomitant generation of accessible chromatin. Deletion of the transactivation domain retains the chromatin binding ability of GATA3 but cripples chromatin reprogramming ability, resulting in failure to induce MET.ConclusionsThese data provide mechanistic insights into GATA3-mediated chromatin reprogramming during MET, and suggest unexpected complexity to TF pioneering. Successful reprogramming requires stable binding to a nucleosomal site; activation domain-dependent recruitment of co-factors including BRG1, the ATPase subunit of the SWI/SNF chromatin remodeling complex; and appropriate genomic context. The resulting model provides a new conceptual framework for de novo enhancer establishment by a pioneer TF.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-016-0897-0) contains supplementary material, which is available to authorized users.

ertilization and early embryogenesis involve the transition from specialized unipotent gametes to totipotent embryos. After fertilization, mammalian embryos rely on maternally deposited RNA but subsequently initiate ZGA during which embryonic transcription begins 1 . Diverse mechanisms control the timing of ZGA, such as controlling RNA polymerase activity, the nuclear/ cytoplasmic ratio, or translation of critical ZGA transcription factors (TFs) in Caenorhabditis elegans, Drosophila melanogaster or Danio rerio, respectively 1 . Currently, we have an incomplete understanding of how the transcriptional machinery (RNA polymerases) and/or sequence-specific TFs dictate the timing of ZGA in mammals and contribute to developmental potential.Recent work identified the TF DUX (DUX4 in humans) as a key regulator of ZGA gene expression 2-5 . When ectopically expressed in cells, DUX and DUX4 activate many ZGA genes, including the earliest wave of ZGA genes in humans and mice 2,3 . However, the extent to which DUX is required for appropriate ZGA is unclear, as the reported effect of genetic loss of Dux ranges from minor molecular to major transcriptional defects and decreased development in mouse or human embryos 4-6 .To study ZGA using a cellular model, we and others have used 2C-embryo-like cells (2CLCs), which are an endogenously fluctuating subpopulation of mouse embryonic stem cells (mESCs) that recapitulate several key features of ZGA 7 . The 2CLCs activate transcripts characteristic of the 2C mouse embryo (including Dux,