Histone variant H2A.X deposition pattern serves as a functional epigenetic mark for distinguishing the developmental potentials of iPSCs.

For future application of induced pluripotent stem cell (iPSC) technology, the ability to assess the overall quality of iPSC clones will be an important issue. Here we show that the histone variant H2A.X is a functional marker that can distinguish the developmental potentials of mouse iPSC lines. We found that H2A.X is specifically targeted to and negatively regulates extraembryonic lineage gene expression in embryonic stem cells (ESCs) and prevents trophectoderm lineage differentiation. ESC-specific H2A.X deposition patterns are faithfully recapitulated in iPSCs that support the development of "all-iPS" animals via tetraploid complementation, the most stringent test available of iPSC quality. In contrast, iPSCs that fail to support all-iPS embryonic development show aberrant H2A.X deposition, upregulation of extraembryonic lineage genes, and a predisposition to extraembryonic differentiation. Thus, our work has highlighted an epigenetic mechanism for maintaining cell lineage commitment in ESCs and iPSCs that can be used to distinguish the quality of iPSC lines.

[1]  Janet Rossant,et al.  Cdx2 is required for correct cell fate specification and differentiation of trophectoderm in the mouse blastocyst , 2005, Development.

[2]  E. Rogakou,et al.  Histone H2A variants H2AX and H2AZ. , 2002, Current opinion in genetics & development.

[3]  K. Hochedlinger,et al.  A reprogrammable mouse strain from gene-targeted embryonic stem cells , 2010, Nature Methods.

[4]  J. Rossant.  Stem cells and lineage development in the mammalian blastocyst. , 2007, Reproduction, fertility, and development.

[5]  Michel C. Nussenzweig,et al.  Genomic Instability in Mice Lacking Histone H2AX , 2002, Science.

[6]  G. Pruneri,et al.  Cell Reprogramming Requires Silencing of a Core Subset of Polycomb Targets , 2013, PLoS genetics.

[7]  Yuriy L Orlov,et al.  Eset partners with Oct4 to restrict extraembryonic trophoblast lineage potential in embryonic stem cells. , 2009, Genes & development.

[8]  R. Young,et al.  SetDB1 contributes to repression of genes encoding developmental regulators and maintenance of ES cell state. , 2009, Genes & development.

[9]  Riitta Lahesmaa,et al.  Tet1 and Tet2 regulate 5-hydroxymethylcytosine production and cell lineage specification in mouse embryonic stem cells. , 2011, Cell stem cell.

[10]  Andrew D. Smith,et al.  Bioinformatics Applications Note Gene Expression Identifying Dispersed Epigenomic Domains from Chip-seq Data , 2022 .

[11]  Yali Chen,et al.  Expression of Fgfr2 in the early mouse embryo indicates its involvement in preimplantation development , 1999, Mechanisms of Development.

[12]  F. P. Villena,et al.  Genome imprinting regulated by the mouse Polycomb group protein Eed , 2003, Nature Genetics.

[13]  Y. Kluger,et al.  Picking ChIP-seq peak detectors for analyzing chromatin modification experiments , 2012, Nucleic acids research.

[14]  Christoph Bock,et al.  Sequential ChIP-bisulfite sequencing enables direct genome-scale investigation of chromatin and DNA methylation cross-talk , 2012, Genome research.

[15]  Janet Rossant,et al.  Gata 3 regulates trophoblast development downstream of Tead 4 and in parallel to Cdx 2 , 2022 .

[16]  J. Rossant,et al.  Gata3 regulates trophoblast development downstream of Tead4 and in parallel to Cdx2 , 2010, Development.

[17]  Rudolf Jaenisch,et al.  Mechanisms and models of somatic cell reprogramming , 2013, Nature Reviews Genetics.

[18]  Panayiotis V. Benos,et al.  STAMP: a web tool for exploring DNA-binding motif similarities , 2007, Nucleic Acids Res..

[19]  E. Davis,et al.  Homologous Recombination, DNA Repair, and Mycobacterial recA Genes , 1994 .

[20]  C. Glass,et al.  Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. , 2010, Molecular cell.

[21]  T. Ludwig,et al.  Homologous recombination DNA repair genes play a critical role in reprogramming to a pluripotent state. , 2013, Cell reports.

[22]  Satoshi Tanaka,et al.  Trophoblast stem cells. , 2006, Methods in enzymology.

[23]  Wei Li,et al.  Activation of the Imprinted Dlk1-Dio3 Region Correlates with Pluripotency Levels of Mouse Stem Cells , 2010, The Journal of Biological Chemistry.

[24]  H. Ueda,et al.  Context-dependent wiring of Sox2 regulatory networks for self-renewal of embryonic and trophoblast stem cells. , 2013, Molecular cell.

[25]  H. Kimura,et al.  Proviral silencing in embryonic stem cells requires the histone methyltransferase ESET , 2010, Nature.

[26]  Radu Dobrin,et al.  Dissecting self-renewal in stem cells with RNA interference , 2006, Nature.

[27]  Janet Rossant,et al.  Interaction between Oct3/4 and Cdx2 Determines Trophectoderm Differentiation , 2005, Cell.

[28]  F. Alt,et al.  Increased ionizing radiation sensitivity and genomic instability in the absence of histone H2AX , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[29]  Ryan A. Flynn,et al.  A unique chromatin signature uncovers early developmental enhancers in humans , 2011, Nature.

[30]  J. Miyazaki,et al.  Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells , 2000, Nature Genetics.

[31]  David A. Williams,et al.  Overcoming reprogramming resistance of Fanconi anemia cells. , 2012, Blood.

[32]  C. Allis,et al.  Histone variants in metazoan development. , 2010, Developmental cell.

[33]  W. Reik,et al.  Epigenetic restriction of embryonic cell lineage fate by methylation of Elf5 , 2008, Nature Cell Biology.

[34]  P. Cahan,et al.  Origins and implications of pluripotent stem cell variability and heterogeneity , 2013, Nature Reviews Molecular Cell Biology.

[35]  Stuart H. Orkin,et al.  Chromatin Connections to Pluripotency and Cellular Reprogramming , 2011, Cell.

[36]  Tomohiro Kono,et al.  Aberrant silencing of imprinted genes on chromosome 12qF1 in mouse induced pluripotent stem cells , 2010, Nature.

[37]  Jarrett Rosenberg,et al.  Single cell transcriptional profiling reveals heterogeneity of human induced pluripotent stem cells. , 2011, The Journal of clinical investigation.

[38]  Geppino Falco,et al.  Uncovering early response of gene regulatory networks in ESCs by systematic induction of transcription factors. , 2009, Cell stem cell.

[39]  Manuel Serrano,et al.  A p53-mediated DNA damage response limits reprogramming to ensure iPS cell genomic integrity , 2009, Nature.

[40]  Kristian Helin,et al.  Genome-wide mapping of Polycomb target genes unravels their roles in cell fate transitions. , 2006, Genes & development.