Revisiting the consequences of deleting the X inactivation center.

Abstract

Mammalian cells equalize X-linked dosages between the male (XY) and female (XX) sexes by silencing one X chromosome in the female sex. This process, known as "X chromosome inactivation" (XCI), requires a master switch within the X inactivation center (<i>Xic</i>). The <i>Xic</i> spans several hundred kilobases in the mouse and includes a number of regulatory noncoding genes that produce functional transcripts. Over three decades, transgenic and deletional analyses have demonstrated both the necessity and sufficiency of the <i>Xic</i> to induce XCI, including the steps of X chromosome counting, choice, and initiation of whole-chromosome silencing. One recent study, however, reported that deleting the noncoding sequences of the <i>Xic</i> surprisingly had no effect for XCI and attributed a sufficiency to drive counting to the coding gene, <i>Rnf12/Rlim</i> Here, we revisit the question by creating independent <i>Xic</i> deletion cell lines. Multiple independent clones carrying heterozygous deletions of the <i>Xic</i> display an inability to up-regulate Xist expression, consistent with a counting defect. This defect is rescued by a second site mutation in <i>Tsix</i> occurring <i>in trans</i>, bypassing the defect in counting. These findings reaffirm the essential nature of noncoding <i>Xic</i> elements for the initiation of XCI.