Implication of the Mediator kinase module in CIZ1 recruitment and gene silencing by <i>Xist</i> during the initiation of X inactivation

In mammals, dosage compensation between the sexes is mediated by X chromosome inactivation. The long non-coding <i>Xist</i> RNA initiates this process. The molecular mechanism of X-linked gene silencing is beginning to be understood. We have previously reported that the cyclin-dependent kinase CDK8 is required for gene silencing and histone H3 trimethyl-lysine 27 recruitment by <i>Xist</i>. CDK8 functions in cell signalling and transcriptional regulation, in particular, it associates with MED12, MED13, and cyclin C to form a kinase submodule of the Mediator complex. It remains unknown if CDK8 acts as a Mediator kinase in X chromosome inactivation. Here, we analyse a <i>Med12</i> mutation in an embryonic stem cell model for the initiation of X chromosome inactivation. Loss of <i>Med12</i> is compatible with self-renewal and survival of mouse embryonic stem cells, but impairs X-linked gene silencing by <i>Xist</i>. In addition, recruitment of CIZ1 and histone H3 trimethyl-lysine 27 by <i>Xist</i> are significantly reduced in the absence of <i>Med12</i>. We further show that CIZ1 recruitment is also modulated by <i>Cdk8</i>. Our study shows that mutations of <i>Med12</i> and <i>Cdk8</i> have similar effects on <i>Xist</i> function and provide a cell system for studying the role of the Mediator kinase module in X chromosome inactivation.