G1/S restriction point controls phasic gene expression during early mouse development and differentiation of ESCs

Pluripotent embryonic stem cells have a unique cell cycle structure with a suppressed G1/S restriction point and little differential expression across the cell cycle phases. Here, we evaluate the link between G1/S restriction point activation, phasic gene expression, and cellular differentiation. Expression analysis reveals a gain in phasic gene expression across lineages between embryonic days E7.5 and E9.5. Genetic manipulation of the G1/S restriction point regulators miR-302 and P27 respectively accelerates or delays the onset of phasic gene expression in mouse embryos. Relief of miR-302-mediated p21 or p27 suppression expedites embryonic stem cell differentiation, while a constitutive Cyclin E mutant blocks it. Together, these findings uncover a causal relationship between emergence of the G1/S restriction point with a gain in phasic gene expression and cellular differentiation. mRNA and miRNA expression profiling between cell cycle phases in bulk and single cell E7.5, E8.5 and E9.5 mouse embryos. Wild-type embryos are profiled in all cases, miR-302 knockout and p27 knockout embryos are profiled in a subset of the experiments. mRNA expression profiles of cell cycle phase sorted 3T3, embryonic stem cell (ESC) and differentiating ESCs (wt and Ccne1 mutant) are also included. Comparative gene expression profiling analysis of RNA-seq data for cell cycle phase sorted 3T3, ESC and differentiated (wild type and mutant) cells. This work was also funded by a CIHR Fellowship awarded to Brian DeVeale.