Preventing erosion of X-chromosome inactivation in human embryonic stem cells

Abstract

X-chromosome inactivation is a paradigm of epigenetic transcriptional regulation. Female human embryonic stem cells (hESCs) often undergo erosion of X-inactivation upon prolonged culture. Here, we investigate the sources of X-inactivation instability by deriving new primed pluripotent hESC lines. We find that culture media composition dramatically influenced the expression of XIST lncRNA, a key regulator of X-inactivation. hESCs cultured in a defined xenofree medium stably maintained XIST RNA expression and coating, whereas hESCs cultured in the widely used mTeSR1 medium lost XIST RNA expression. We pinpointed lithium chloride in mTeSR1 as a cause of XIST RNA loss. The addition of lithium chloride or inhibitors of GSK-3 proteins that are targeted by lithium to the defined hESC culture medium impeded XIST RNA expression. GSK-3 inhibition in differentiating female mouse embryonic stem cells and epiblast stem cells also resulted in a loss of XIST RNA expression. Together, these data may reconcile observed variations in X-inactivation in hESCs and inform the faithful culture of pluripotent stem cells.

Lineage-specific dynamics of erasure of X-upregulation during inactive-X reactivation

“…X-linked genes play a vital role in different biological pathways, including early development, cell fate specification and pluripotency exit (Cloutier et al, 2022; Marahrens et al, 1997; Schulz et al, 2014; Zhang et al, 2016). However, X-linked genes in male and female cells are expressed monoallelically.…”

Attempts for deriving extended pluripotent stem cells from common marmoset embryonic stem cells

“…We note that the X‐chromosome inactivation state in human EPSCs has not been scrutinized (Yang, Liu, et al, 2017). More recently, Cloutier et al (2022) pinpointed the evolutional difference in X‐chromosome inactivation in vivo between mice and humans (Cloutier et al, 2022). They showed that the X‐chromosome inactivation in vitro can be controlled by lithium ion.…”

Induction of fetal meiotic oocytes from embryonic stem cells in cynomolgus monkeys

“…Although it has been shown that DNA methylation of the XIST promoter on Xi is involved in Xi erosion (Fukuda et al , 2021), the detailed mechanism underlying Xi erosion remains unclear. Notably, however, a recent study has shown that lithium chloride (LiCl), a component of conventional hPSC culture media such as mTeSR1, activates WNT signaling and induces Xi erosion, and the removal of LiCl from the medium prevents hPSCs from Xi erosion (Cloutier et al , 2022). LiCl also induces Xi erosion in mouse epiblast stem cells (EpiSCs; Cloutier et al , 2022).…”