Methylation of the Human AR Locus Does Not Correlate with the Presence of Inactivated X Chromosome in Induced Pluripotent Stem Cells

The process of dosage compensation of genes occurs in the female mammalian cells during early embryonic development: one of the two X chromosomes becomes transcriptionally inactive and is subsequently inherited in the inactive state in all somatic cells. Embryonic stem cells (ESCs) together with induced pluripotent stem cells (iPSCs) have become the models for studying early development. In human female iPSCs and ESCs, one of the X chromosomes is always active, while the state of the other X chromosome can vary: it can be completely active or partially or completely inactivated as in somatic cells. Determining the state of inactivation of the X chromosome in human pluripotent stem cells (PSC) appears to be a complicated and state-of the-art question. However, various methods for determining the inactivation state frequently result in the contradictory findings. The analysis of the AR gene methylation, which is widely used in clinical laboratories, makes it possible to determine the ratio of alleles of the inactivated X chromosome in a patient’s blood cells and tissues. Previously, this approach was suggested to determine the status of X chromosome inactivation in human iPSCs. In the present study, we analyzed the inactivation state of the X chromosome in the human iPSC line by standard methods (XIST gene expression, XIST promoter methylation, histone modification, etc.), as well as by the level of methylation of the AR gene via fragment analysis. The methylation of the AR locus failed to correlate with the X chromosome inactivation state in female iPSCs. Thus, we assume that the methylation of the AR gene cannot be used as an indicator of X chromosome inactivation in human female iPSC lines.