Human Naive Pluripotent Stem Cells Model X Chromosome Dampening and X Inactivation

Abstract

Summary Naïve human embryonic stem cells (hESCs) can be derived from primed hESCs or directly from blastocysts, but their X-chromosome state has remained unresolved. Here, we show that the inactive X-chromosome (Xi) of primed hESCs was reactivated in naïve culture conditions. Like cells of the blastocyst, the resulting naive cells contained two active X-chromosomes with XIST expression and chromosome-wide transcriptional dampening, and initiated XIST-mediated X-inactivation upon differentiation. Both establishment of and exit from the naïve state (differentiation) happened via an XIST-negative XaXa intermediate. Together, these findings identify a cell culture system for functionally exploring the two X-chromosome dosage compensation processes in early human development: X-dampening and X-inactivation. However, remaining differences between naïve hESCs and embryonic cells related to monoallelic XIST expression and non-random X-inactivation highlight the need for further culture improvement. As the naïve state reset Xi abnormalities seen in primed hESCs it may provide cells better suited for downstream applications.

When the Lyon(ized chromosome) roars: ongoing expression from an inactive X chromosome

“…Nevertheless, evaluation of escapees at the onset of human XCI has been hampered, as human ESCs generally retain an inactive X chromosome that erodes upon culturing, which has limited the study of early human XCI in vitro. However, recent work suggests that human naïve ESC can be isolated that model the earliest steps of human XCI [113], and thus it may be possible to determine whether human escapees are ever silenced.…”

Epigenetic foundations of pluripotent stem cells that recapitulate in vivo pluripotency

“…83 Consistent with ICM-like pluripotency, human ES cells in these conditions display global DNA hypomethylation and the gene expression patterns of naive markers and transposons, which are reminiscent of preimplantation human embryos [84][85][86] ( Figure 5). Of particularly note, inactive X chromosome becomes active in naive-like female cells in vitro, suggesting that reactivation of X chromosome occurs during naive conversion 84,87,88 (Figure 5). Therefore, it is expected that human naive cells serve as a powerful tool for studying human early embryogenesis and human diseases.…”

Naive Pluripotent Stem Cells as a Model for Studying Human Developmental Epigenomics: Opportunities and Limitations

“…Another defining characteristic of naive hESCs is the presence of two active X chromosomes in female cells [13,22,23]. This discovery is particularly exciting because it presents a major difference from conventional hESCs, which typically have one active and one inactive X-chromosome (reflecting their postimplantation status).…”

“…The presence of two active X chromosomes is stable in naive hESCs, and most cells co-express XIST and XACT. Interestingly, the proportion of cells with biallelic XIST transcription varies between cell lines and over passage [22,23], and it is unclear why this expression pattern should be unstable or potentially detrimental.…”

“…One X-chromosome is inactivated upon naive hESC differentiation, although curiously the choice of which chromosome is inactivated appears to be nonrandom [13,23]. Notably, the two studies that showed this observation used naive hESCs that were reprogrammed from conventional hESCs, and so this unanticipated observation could be due to an epigenetic 'memory' of the previously active chromosome.…”