Spontaneous reactivation of the inactive X chromosome in mouse embryonal carcinoma cells

Abstract

The mouse embryonal carcinoma cell line MC12 carries two X chromosomes, one of which replicates late in S phase and shares properties with the normal inactive X chromosome and, therefore, is considered to be inactivated. Since the hypoxanthine phosphoribosyl transferase (HPRT) gene on the active X chromosome is mutated (Hprt^–), MC12 cells lack HPRT activity. After subjecting MC12 cells to selection in HAT medium, however, a number of HAT-resistant clones (HAT^R) appeared. The high frequency of HAT resistance (3.18 × 10^–4) suggested reactivation of Hprt⁺ on the inactive X chromosome rather than reversion of Hprt^–. Consistent with this view, cytological analyses showed that the reactivation occurred over the length of the inactive X chromosome in 11 of 20 HAT^R clones isolated. The remaining nine clones retained a normal heterochromatic inactive X chromosome. The spontaneous reactivation rate of the Hprt⁺ on the inactive X chromosome was relatively high (1.34 × 10^–6) and comparable to that observed for Xist-deleted somatic cells (Csankovszki et al., 2001), suggesting that the inactivated state is poorly maintained in MC12 cells.