GSE98771
Continuous suppression of Mek1/2 impairs the developmental potential of mouse embryonic stem cells (H2A.X pattern)
Simultaneous inhibition of Gsk3a/ß and Mek1/2 signaling in the presence of LIF (2i/L) induces a naïve state in mouse embryonic stem cells (ESCs) that resembles the inner cell mass (ICM) of the pre-implantation embryo. Since the ICM exists only transiently in vivo, it remains unclear how continuous propagation of naïve ESCs in vitro affects their stability and functionality. Here, we show that extended culture of male ESCs in 2i/L results in the progressive erosion of genomic imprints, loss of H2A.X binding, and accumulation of chromosomal aberrations. Consistent with these observations, we find that ESCs propagated in 2i/L have an impaired developmental potential. Mechanistically, we demonstrate that Mek1/2 inhibition is predominantly responsible for these effects, in part through downregulation of DNA methyltransferases. We further provide evidence that female ESCs cultured in conventional serum/LIF media phenocopy male ESCs cultured in 2i/L, including the aforementioned epigenetic and developmental abnormalities. Finally, we show that replacement of the Mek1/2 inhibitor with a Src inhibitor preserves the epigenetic and genomic integrity as well as developmental potential of ESCs. Taken together, our data suggest that, while suppression of Mek1/2 in ESCs maintains an ICM-like epigenetic state, continuous suppression results in irreversible changes that compromise their developmental potential. Overall design: Detect and analyze different H2A.X deposition pattern in ESCs under different culture conditions