Maintenance of X inactivation of theRps4, Zfx, andUbe1 genes in a mouse in vitro system

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Abstract
Several genes, includingRPS4X (ribosomal protein subunit 4),ZFX (zinc finger on the X chromosome), andUBE1 (ubiquitin-activating enzyme), have been shown to be expressed from the inactive X chromosome of cultured human cells. By contrast, these genes are subject to X-chromosome inactivation in tissues from adult mice. We have now examined the inactivation status of these genes in cultured mouse cells to determine whether the differences in X-chromosome inactivation between species is due to an intrinsic difference between human and mouse X-chromosome genes or whether it is a function of gene reactivation in cell culture per se. The expression of three mouse X-chromosome genes,Rps4, Zfx, andUbe1 was examined by reverse transcriptase polymerase chain reaction (RT-PCR) in heterozygous cultured cells from a cross of a laboratory mouse byMus spretus, which were selected to uniformly express the X chromosome from the laboratory mouse parent. No expression of theM. spretus alleles of these genes was observed in the cell line (Hobmski), which is consistent with the patterns of expression previously observed in mouse in vivo and indicates that these genes remain stably inactivated in an immortalized mouse cell line. By cytogenetic and RT-PCR analyses the Hobmski cell line was shown to retain a late-replicating X chromosome fromM. spretus, which expressed theM. spretus allele of the X (inactive) specific transcript (Xist). The Hobmski cell line will be a useful resource for studying the features that maintain X-chromosome genes inactive.