Changes in the position and volume of inactive X chromosomes during the G0/G1 transition

In female mammals, each cell silences one X chromosome by converting it into transcriptionally inert heterochromatin. The inactivation is concomitant with epigenetic changes including methylation of specific histone residues and incorporation of macroH2A. Such epigenetic changes may exert influence on the positioning of the inactive X chromosome (Xi) within the nucleus beyond the level of chromatin structure. However, the dynamic positioning of the inactive X chromosome during cell cycle remains unclear. Here, we show that H3K27me3 is a cell-cycle-independent marker for the inactivated X chromosomes in WI38 cells. By utilizing this marker, three types of Xi locations in the nuclei are classified, which are envelope position (associated with envelope), mid-position (between the envelope and nucleolus), and nucleolus position (associated with the nucleolus). Moreover, serial-section analysis revealed that the inactive X chromosomes in the mid-position appear to be sparser and less condensed than those associated with the nuclear envelope or nucleolus. During the transition from G0 to G1 phase, the inactive X chromosomes tend to move from the envelope position to the nucleolus position in WI38 cells. Our results imply a role of chromosome positioning in maintaining the organization of the inactive X chromosomes in different cell phases.