Pou5f3 and Nanog organize active chromatin in three dimensions

Zygotic genome in zebrafish embryogenesis is activated by transcription factors Pou5f3, Sox19b and Nanog. While it was previously demonstrated, that these factors displace nucleosomes and create the regions of open chromatin, their role in three-dimensional genome organisation was not studied. Here we show, that large-scale chromatin structures, TADs and compartments, form independently of Pou5f3, Sox19b and Nanog. However, Pou5f3 and Nanog are required for formation of small-scale 3D active chromatin structures which we call "fountains". Overall design: To investigate if Pou5f3, Sox19b and Nanog play a role in 3D organization of chromatin , we compared 3D chromatin maps using Hi-C method in the wild-type, single, double and triple mutants by these factors. Hi-C was performed shortly before zygotic genome activation ( 2.75 hours post-fertilization, or hpf), and after zygotic genome activation, at late blastula stage ( 5.3 hpf). To characterize the developmental changes of 3D chromatin , we also performed Hi-C on the wild-type at 11 hpf and 25 hpf.