A SAGE approach to identifying novel <i>trans</i>-acting factors involved in the X inactivation process

Abstract

X chromosome inactivation ensures the dosage compensation of X-linked genes in XX females compared to their XY male counterpart. It is characterised by the specific recruitment of an inhibitory ribonucleoprotein complex involving the non-coding Xist RNA to the presumptive inactive X chromosome and associated chromatin modifications, which result in the transcriptional silencing of the X chromosome. As an approach to the identification of some of the potential molecular players in this process we have performed comparative transcriptional profiling of mouse 6.5-dpc (days post-coitum) female and male embryos using a modified SAGE (Serial analysis of gene expression) technique which allows the analysis of small quantities of biological material. At 6.5 dpc, a moment when random X inactivation of embryonic tissues has just been achieved, some two hundred transcripts that were significantly enriched in the female gastrula compared to its male counterpart could be identified. The validation of an association with the X inactivation process of a subset of these transcripts has been studied, ex vivo, in differentiating female and male ES cells and in female ES cells in which the establishment of X inactivation is interrupted through the post-transcriptional inhibition of Xist synthesis.

Nuclear mRNA Degradation Pathway(s) Are Implicated in Xist Regulation and X Chromosome Inactivation

“…We reasoned that if there were differences in transcript frequencies between female and male embryos at this stage, a subset of such differences might be implicated in X-inactivation. More than 214 genes overexpressed in female compared with male were identified [ 13 ], and a subset was validated using the ES cell system. Of the candidates falling within the category of upregulated genes and which are implicated in RNA metabolism and processing, the Eif1 (other name, Sui1-rs1 ) gene was retained for functional analysis.…”