Identifying Reprogramming Barriers to Totipotency

The transition between totipotency and pluripotency provides an excellent model for studying cellular plasticity and cell fate specification in mammals. Totipotent cells can generate both embryonic and extraembryonic tissues and appear transiently as the embryo progresses from a single-cell zygote to the multicellular blastocyst. The blastocyst contains the inner cell mass (ICM), whose potential, in contrast to totipotent cells, is restricted to embryonic tissues. However, the ICM-derived embryonic stem cells or the functionally equivalent induced pluripotent stem cells (iPSCs) contain a rare population of 2-cell embryo-like cells (2CLCs) that display expanded 2CLC potency and cycle in-and-out of the pluripotent state. Morf4l1 is a highly conserved epigenetic regulator involved in direct reprogramming. However, whether Morf4l1 contributes to reprogramming pluripotent stem cells to totipotent cells remains to be determined. Here we show that Morf4l1 represses the induction of 2CLC in mouse iPSC culture. Loss of Morf4l1 induces an intermediate cell type with some transcriptome and chromatin features similar to 4-cell and 8-cell mouse embryos. Specifically, Morf4l1 contributes to the regulation of nucleolar morphology, rRNA biogenesis, and chromatin accessibility as cells transit between pluripotent and 2CLC states. Furthermore, loss- and gain-of-function studies indicate that Morf4l1 is an integral component of an epigenetic program that facilitates the 2C-like transition. This study establishes Morf4l1 as a critical barrier in a mammalian culture system of early cell fate specification and offers a non-chemical-mediated strategy to enhance 2C-like reprogramming.