title: Aberrant patterns of X chromosome inactivation in a new line of human embryonic stem cells established in physiological oxygen concentrations.
abstract: 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.
authors.affiliation: Laboratory of Molecular Genetics, University of São Paulo, São Paulo, Brazil.
authors.forename: Juliana Andrea
authors.initials: JA
authors.lastname: de Oliveira Georges
authors.forename: Naja
authors.initials: N
authors.lastname: Vergani
authors.forename: Simone Aparecida Siqueira
authors.initials: SA
authors.lastname: Fonseca
authors.forename: Ana Maria
authors.initials: AM
authors.lastname: Fraga
authors.forename: Joana Carvalho Moreira
authors.initials: JC
authors.lastname: de Mello
authors.forename: Maria Cecília R Maciel
authors.initials: MC
authors.lastname: Albuquerque
authors.forename: Litsuko Shimabukuro
authors.initials: LS
authors.lastname: Fujihara
authors.forename: Lygia Veiga
authors.initials: LV
authors.lastname: Pereira
mesh_terms: D000818:Animals; D002454:Cell Differentiation; D002478:Cells, Cultured; D041321:Chromosomes, Human, X; D020732:Cytogenetic Analysis; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism*; D005260:Female; D006657:Histones / Q000378:metabolism; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D016513:Mice, SCID; D010100:Oxygen / Q000378:metabolism*; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D062085:RNA, Long Noncoding / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013724:Teratoma / Q000378:metabolism / Q000473:pathology; D049951:X Chromosome Inactivation
chemical_list: D006657:Histones; D062085:RNA, Long Noncoding; D012333:RNA, Messenger; C077885:XIST non-coding RNA; D010100:Oxygen