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

• Published in: Nature communications Vol. 13; no. 1; pp. 2516 - 18
• Main Authors: Cloutier, Marissa, Kumar, Surinder, Buttigieg, Emily, Keller, Laura, Lee, Brandon, Williams, Aaron, Mojica-Perez, Sandra, Erliandri, Indri, Rocha, Andre Monteiro Da, Cadigan, Kenneth, Smith, Gary D., Kalantry, Sundeep
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 06.05.2022; Nature Publishing Group; Nature Portfolio
• Subjects: 14; 14/32; 45; 45/91; 631/208/200; 631/532/2117; 631/532/2442; Animals; Cell culture; Cell differentiation; Chromosomes; Chromosomes - metabolism; Culture media; Deactivation; Embryo cells; Epigenetics; Erosion control; Female; Females; Gene expression; Gene regulation; Glycogen Synthase Kinase 3 - metabolism; Human Embryonic Stem Cells - metabolism; Humanities and Social Sciences; Humans; Inactivation; Inhibitors; Lithium; Lithium chloride; Lithium Chloride - metabolism; Mice; multidisciplinary; Pluripotency; Proteins; Ribonucleic acid; RNA; RNA, Long Noncoding - genetics; RNA, Long Noncoding - metabolism; Science; Science (multidisciplinary); Stem cell transplantation; Stem cells; Transcription; X Chromosome Inactivation; X chromosomes
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-022-30259-x

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. Cloutier et al. discover that human embryonic stem cells (hESCs) cultured with media containing inhibitors of GSK3 proteins undergo erosion of X-chromosome inactivation, which equalizes X-linked gene expression between females and males. The findings inform the faithful culture of hESCs.