Characterization of Xpr (Xpct) reveals instability but no effects on X-chromosome pairing or Xist expression

X-chromosome inactivation balances X-chromosome dosages in male and female mammals by transcriptionally repressing one X in the female sex. Proper counting and the mutually exclusive choice of active X and inactive X have been hypothesized to involve X-chromosome crosstalk via homologous chromosome pairing. Transient pairing of two female Xs requires noncoding Tsix and Xite. A recent study suggested a new pairing element (Xpr), located ~200 kb upstream of Xist, in the Xpct region. Xpr is proposed to induce pairing and activate Xist expression. Here, we further characterize Xpr and find that the Xpr sequence is unstable when introduced as transgenes into male ES cells. Xpr transgenes show an unusual tendency to disperse throughout the nucleus. However, we observe neither pairing between Xpr alleles nor ectopic Xist expression. In the absence of Tsix, Xpr does not induce inter-allelic Xic interactions. Female ES cells carrying Xpr transgenes are more stable. Nonetheless, pairing also does not seem to occur in female cells. We conclude that, while Xpr contains unusual properties, it most likely does not serve as a pairing or counting element. Differences in statistical methods and controls may explain some of the discrepancies.