PLX067101

GSE116413: Unfolding the inactive X chromosome

• Organsim mouse
• Type RNASEQ
• Target gene
• Project ARCHS4

The inactive X chromosome (Xi) is folded in a stepwise process into a compartment-less structure. Here we investigate if the Xi can be unfolded in post-XCI cells. We began with ablating structural maintenance of chromosomes hinge domain containing 1 (Smchd1) in female mouse embryonic fibroblasts (MEFs), a cell type in which XCI is completed. We then performed in situ Hi-C on one Smchd1+/+ (wild-type, WT) and one Smchd1-/- (knockout, KO) MEF clone to probe the role of SMCHD1 in maintaining the higher-order structure of the Xi. To determine if the change in chromosome structure is accompanied by altered gene expression, we performed RNA-seq on two WT and two independently generated Smchd1-/- clones treated with either DMSO or 5-aza-2'-deoxycytidine (Aza), a DNA demethylating agent. In addition, we performed H3K27me3 and H2AK119ub ChIP-seq and Xist Capture Hybridization Analysis of RNA Targets with deep sequencing (CHART-seq) on one WT and one Smchd1-/- MEF clone to determine if there is an alteration in the epigenetic state of the Smchd1-/- Xi. Finally, we examined the role of Xist, Polycomb repressive complex 1 (PRC1), and Heterogeneous Nuclear Ribonucleoprotein K (HNRNPK) in regulating the Xi structure in post-XCI cells. To do so we performed in situ Hi-C on WT and Smchd1-/- MEFs depleted with either PRC1 or HNRNPK, as well as on fibroblasts in which Xist is deleted on the Xi (XidelXist). SOURCE: Chen-Yu Wang (cywang@molbio.mgh.harvard.edu) - Jeannie T. Lee Massachusetts General Hospital