Abstract
Mouse embryonic stem cells (mESCs) cycle in and out of a transient 2-cell (2C)-like totipotent state, driven by a complex genetic circuit involves both the coding and repetitive sections of the genome. While a vast array of regulators, including the multi-functional protein Rif1, has been reported to influence the switch of fate potential, how they act in concert to achieve this cellular plasticity remains elusive. Here, by modularizing the known totipotency regulatory factors, we identify an unprecedented functional connection between Rif1 and the non-canonical polycomb repressive complex PRC1.6. Downregulation of the expression of either Rif1 or PRC1.6 subunits imposes similar impacts on the transcriptome of mESCs. The LacO-LacI induced ectopic colocalization assay detects a specific interaction between Rif1 and Pcgf6, bolstering the intactness of the PRC1.6 complex. Chromatin immunoprecipitation followed by sequencing (ChIP-seq) analysis further reveals that Rif1 is required for the accurate targeting of Pcgf6 to a group of genomic loci encompassing many genes involved in the regulation of the 2C-like state. Depletion of Rif1 or Pcgf6 not only activates 2C genes such as Zscan4 and Zfp352, but also derepresses a group of the endogenous retroviral element MERVL, a key marker for totipotency. Collectively, our findings discover that Rif1 can serve as a novel auxiliary component in the PRC1.6 complex to restrain the genetic circuit underlying totipotent fate potential, shedding new mechanistic insights into its function in regulating the cellular plasticity of embryonic stem cells.