Capture of Mouse and Human Stem Cells with Features of Formative Pluripotency

Abstract

Summary Pluripotent cells emerge as a naive founder population in the blastocyst, acquire capacity for germline and soma formation, and then undergo lineage priming. Mouse embryonic stem cells (ESCs) and epiblast-derived stem cells (EpiSCs) represent the initial naive and final primed phases of pluripotency, respectively. Here, we investigate the intermediate formative stage. Using minimal exposure to specification cues, we derive stem cells from formative mouse epiblast. Unlike ESCs or EpiSCs, formative stem (FS) cells respond directly to germ cell induction. They colonize somatic tissues and germline in chimeras. Whole-transcriptome analyses show similarity to pre-gastrulation formative epiblast. Signal responsiveness and chromatin accessibility features reflect lineage capacitation. Furthermore, FS cells show distinct transcription factor dependencies, relying critically on Otx2. Finally, FS cell culture conditions applied to human naive cells or embryos support expansion of similar stem cells, consistent with a conserved staging post on the trajectory of mammalian pluripotency.

The ETS Transcription Factor ERF controls the exit from the naïve pluripotent state

“…FA-RAS KO ESC showed high expression of OTX2, which has been shown a critical transcription factor for thew maintenance of intermediate states of pluripotency (7). The consequence of having high levels of OTX2 in this context can be inferred by previous studies (25).…”

“…Finally, we showed that FA-RAS KO ESC can be maintained in culture under primed conditions and could be a suitable genetic model to study intermediate states of pluripotency. Indeed, FA-RAS KO ESC mirrored transcriptionally cells that have departed 12-24 hours from naïve pluripotency and are reminiscent of the recently described pluripotent intermediate states (6)(7)(8). Future studies will be necessary to determine the similarity of all these intermediate states with corresponding in vivo counterparts.…”

“…Inhibition of WNT signaling drives the pluripotent transition from naïve to the rosette state while further activation of the RAS/MAPK signaling promotes the progression to primed pluripotency (6). Furthermore, additional states of intermediate pluripotency characterized by germ cell specification capacity have also been described, formative stem cells (FSC) and chimera pluripotent stem cells (XPSC) (7,8). These intermediates rely on exogenous (XPSC) or autocrine (FSC) FGF signaling for their self-renewal in contrast to the requirements for RSC.…”

“…OTX2 expression is independent of MEK signals but is efficiently repressed by the WNT pathway, thus explaining the elevated levels of OTX2 in rosette-like ESC (6). High levels of OTX2 have also been found to be associated to formative pluripotency (7,8). In addition, we also detected similar high OTX2 levels in FA-RAS KO ESC compared to FA-RAS lox/lox (Fig.…”

Developments in pluripotency: a new formative state

“…In 2020, three other groups have reported the pioneering of 2D culture conditions which support stable in vitro derivation and maintenance of intermediate pluripotent embryonic states. 3,8,9 All these states show cell polarity, a transcriptome indicative of formative pluripotency and, as far as assayed, 8 glycolytic metabolism. Surprisingly, comparative analysis of the transcriptomic and epigenomic data generated by these groups 3,8,9 and in the current study 6 show that the intermediate pluripotent ESCs in each of the culture conditions resemble different time points in the naïve-to-primed transition, over a timespan ranging from approximately E5.0 to E6.5 6 (Fig.…”