Aberrant Patterns of X Chromosome Inactivation in a New Line of Human Embryonic Stem Cells Established in Physiological Oxygen Concentrations
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One of the differences between murine and human embryonic stem cells (ESCs) is the epigenetic state of the X chromosomes in female lines. Murine ESCs (mESCs) present two transcriptionally active Xs that will undergo the dosage compensation process of XCI upon differentiation, whereas most human ESCs (hESCs) spontaneously inactivate one X while keeping their pluripotency. Whether this reflects differences in embryonic development of mice and humans, or distinct culture requirements for the two kinds of pluripotent cells is not known. Recently it has been shown that hESCs established in physiological oxygen levels are in a stable pre-XCI state equivalent to that of mESCs, suggesting that culture in low oxygen concentration is enough to preserve that epigenetic state of the X chromosomes. Here we describe the establishment of two new lines of hESCs under physiological oxygen level and the characterization of the XCI state in the 46,XX line BR-5. We show that a fraction of undifferentiated cells present XIST RNA accumulation and single H3K27me foci, characteristic of the inactive X. Moreover, analysis of allele specific gene expression suggests that pluripotent BR-5 cells present completely skewed XCI. Our data indicate that physiological levels of oxygen are not sufficient for the stabilization of the pre-XCI state in hESCs.
KeywordsHuman embryonic stem cells X chromosome inactivation Hypoxia Epigenetics
This work was supported by grants from Conselho Nacional de Desenvolvimento Científico e Tecnológico/Departamento de Ciência e Tecnologia doMinistério da Saúde (CNPq/MS/DECIT), Banco Nacional de Desenvolvimento Econômico e Social (BNDES), Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) (grant CEPID 2013/08135-2) and Financiadora de Estudos e Projetos (FINEP). AMF and JM have fellowships from FAPESP; NJ and SASF have fellowships from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES).
Conflict of Interest
The authors declare no conflicts of interest.
- 5.Morey, C., & Avner, P. (2011). The demoiselle of X-inactivation: 50 years old and as trendy and mesmerising as ever. PLoS Genetics, 7(7), e1002212. Available: http://www.plosgenetics.org/article/info%3Adoi%2F10.1371%2Fjournal.pgen.1002212. Accessed 06 May 2013.PubMedCentralPubMedCrossRefGoogle Scholar
- 11.Shen, Y., Matsuno, Y., Fouse, S. D., Rao, N., Root, S., et al. (2008). X-inactivation in female human embryonic stem cells is in a nonrandom pattern and prone to epigenetic alterations. Proceedings of the National Academy of Sciences of the United States of America, 105, 4709–4714.PubMedCentralPubMedGoogle Scholar
- 17.Gosden, C., Davdson, C., & Robertson, M. (1992). Lymphocyte culture. In D. E. Rooney & B. H. Czepulkowsky (Eds.), Human cytogenetics (pp. 37–47). Oxford: Oxford University Press.Google Scholar
- 21.Fraga, A. M., Sukoyan, M., Rajan, P., Braga, D. P., Iaconelli, A., Jr., et al. (2011). Establishment of a Brazilian line of human embryonic stem cells in defined medium: implications for cell therapy in an ethnically diverse population. Cell Transplantation, 20(3), 431–440.PubMedCrossRefGoogle Scholar