Analysis of genome architecture during SCNT reveals a role of cohesin in impeding minor ZGA

Somatic cell nuclear transfer (SCNT) can reprogram a somatic nucleus to a totipotent state. However, the re-organization of three-dimensional chromatin structure in this process remains poorly understood. Using low-input Hi-C, we revealed that during SCNT, the transferred nucleus first enters a mitotic-like state (premature chromatin condensation). Unlike fertilized embryos, SCNT embryos show stronger TADs at the 1-cell stage. TADs become weaker at the 2-cell stage, followed by gradual consolidation. Compartments A/B are markedly weak in 1-cell SCNT embryos and become increasingly strengthened afterward. By the 8-cell stage, somatic chromatin architecture is largely reset to embryonic patterns. Unexpectedly, we found cohesin represses minor zygotic genome activation (ZGA) genes (2-cell specific genes) in pluripotent and differentiated cells, and pre-depleting cohesin in donor cells facilitates minor ZGA and SCNT. These data reveal multi-step reprogramming of 3D chromatin architecture during SCNT and support dual roles of cohesin in TAD formation and minor ZGA repression. Overall design: Donor MEF/cumulus cells (MEF/CC), the SCNT embryos at the PCC stage (premature chromosome condensation) (1 hour after nuclear transfer) and SCNT embryos after PCC exit through “SCNT activation” using strontium chloride (SrCl2) at the 1-cell(4hours after activation, 6hours after activation and 10hours after activation), late 2-cell (28 hours after activation), and 8-cell (56 hours after activation) stages were obtained. Both MEF and CC are used for donor cells for the collection of SCNT 8-cell samples. HiC and RNA-seq were performed in these cells at various stages during SCNT embryogenesis. For the predepletion of CTCF/cohesin experiment, the short term differentiated cells with/without auxin treatment are collected for HiC experiments, RNA-seq and ChIP-seq experiments.