Replication Timing Reveal An Epigenetic Commitment To Differentiation Prior To Germ Layer Specification (WG_CGH, RT)

Differentiation of mouse embryonic stem cells (mESCs) is accompanied by global changes in replication timing. To elucidate this reorganization process and explore its potential impact on mouse development, we constructed genome-wide replication-timing profiles of 15 independent mouse cell types representing nine different stages of early mouse development, including all three germ layers. Overall, 45% of the genome exhibits significant changes in replication timing between cell types, indicating that replication-timing regulation is more extensive than previously estimated from neural differentiation. Intriguingly, analysis of early and late epiblast cell culture models suggest that the earliest changes in development include extensive lineage-independent early-to-late replication switches that are completed at a stage equivalent to the post-implantation epiblast, prior to germ layer specification and down-regulation of key pluripotency transcription factors (Oct4/Nanog/Sox2). These changes were stable in all subsequent lineages and involved a class of irreversibly silenced genes that were re-positioned closer to the nuclear periphery. Lineage-specific, late-to-early and early-to-late replication switches followed, which created cell-type specific replication profiles. Importantly, partially reprogrammed induced pluripotent stem cells (piPSCs) failed to restore ESC-specific replication timing and transcription programs particularly within regions of lineage-independent early-to-late replication changes, as well as the inactive X-chromosome. We conclude that lineage-independent, early-to-late replication-timing switches that occur in the post-implantation epiblast embody an epigenetic commitment to differentiation prior to germ layer specification. 22 cell lines, with a total of 36 individual replicates (i.e. 14 in duplicates, 8 in single replicates)