The long noncoding RNA Xist is expressed from only the paternal X chromosome in mouse preimplantation female embryos and mediates transcriptional silencing of that chromosome. In females, absence of Xist leads to postimplantation lethality. Here, through single-cell RNA sequencing of early preimplantation mouse embryos, we found that the initiation of imprinted X-chromosome inactivation absolutely requires Xist. Lack of paternal Xist leads to genome-wide transcriptional misregulation in the early blastocyst and to failure to activate the extraembryonic pathway that is essential for postimplantation development. We also demonstrate that the expression dynamics of X-linked genes depends on the strain and parent of origin as well as on the location along the X chromosome, particularly at the first 'entry' sites of Xist. This study demonstrates that dosage-compensation failure has an effect as early as the blastocyst stage and reveals genetic and epigenetic contributions to orchestrating transcriptional silencing of the X chromosome during early embryogenesis.
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Gene Expression Omnibus
We thank S. Bao and N. Grabole for experimental assistance in single-blastomere RNA sequencing and M. Guttman for sharing the coordinates of Xist entry sites. We are grateful to P. Gestraud and V. Sibut for help in statistical and IPA pathway analysis, respectively. We thank the pathogen-free barrier animal facility of the Institut Curie and J. Iranzo for help with the animals, and the Cell and Tissue Imaging Platform (PICT-IBiSA; member of France–Bioimaging) of the Genetics and Developmental Biology Department (UMR3215/U934) of the Institut Curie for assistance with light microscopy. We acknowledge E. Schulz, E. Nora, I. Okamoto and the members of E.H.'s laboratory for assistance, feedback and critical input. This work was funded by a fellowship from the Région Ile-de-France (DIM STEMPOLE) to M.B.; the Paris Alliance of Cancer Research Institutes (PACRI-ANR) to L.S.; an ERC Advanced Investigator award (ERC-2010-AdG, no. 250367), EU FP7 grants SYBOSS (EU 7th Framework G.A., no. 242129) and MODHEP (EU 7th Framework G.A., no. 259743), La Ligue, Fondation de France, Labex DEEP (ANR-11-LBX-0044), part of the IDEX Idex PSL (ANR-10-IDEX-0001-02 PSL), and ABS4NGS (ANR-11-BINF-0001) to E.H.; France Genomique National infrastructure (ANR-10-INBS-09) to E.H., N.S. and E.B; and CELLECTCHIP (ANR-14-CE10-0013) to E.H. and M.B.
Integrated supplementary information
Summary of single cell RNA-seq samples. For each library isprovided: single cell's name, stage, embryo number, gender, crossand the raw read number, filtered ones and percentage of mapping.
Misregulated genes in absence of paternal Xist. Table summarizing all the genes significantly different between CB wtand CB XistpatΔ female embryos during early development (8-cell toblastocyst).
Pathways and candidate genes downregulated in XistpatΔ femaleblastocysts and highlighted by IPA analysis. Summary of all the pathway and candidate genes and relationships, which have emerged from IPA analysis between CB wt and CB XistpatΔ female blastocysts.