Molecular mechanism of natural pluripotency establishment in the early mouse embryo based on single-cell gene expression profile
Final Report Abstract
Understanding of the molecular mechanism leading to establishment of natural pluripotency in vivo is crucial for the possible application of stem cell biology to regenerative medicine. The aim of this project was to understand the molecular program underlying formation of the pluripotent inner cell mass cells in the mouse blastocyst, which will also elucidate molecular properties of embryonic stem cells. To establish a map of epiblast (EPI) versus primitive endoderm (PrE) lineage segregation which occurs within the inner cell mass (ICM), we comprehensively characterised the gene expression profiles of individual inner cells during blastocyst development. Clustering analysis of the transcriptomes of 66 single inner cells isolated from embryonic day (E)3.25 to E4.5 blastocysts demonstrated that initially they are non-distinguishable. As EPI and PrE lineages become progressively segregated, lineage markers are expressed, but exhibit no apparent correlation early in the segregation process. A hierarchical relationship of lineage-specific marker expression is established only in the late blastocyst. We noted that Fgf4, a key molecule for ICM lineage choice, is differentially expressed at the earliest stage in the segregation, and in its absence the differentiation of ICM cells is halted, indicating that Fgf4 drives, and is required for, ICM lineage segregation. These data led us to propose a model where cell-to-cell heterogeneity generated by stochastic activation of gene expression followed by signal reinforcement underlies ICM lineage segregation by antagonistically separating initially equivalent cells. Future studies that combine the single-cell gene expression analysis and live-imaging of lineage reporter lines will evaluate this model and in particular the possible significance of cell-to-cell heterogeneity in the formation and differentiation of pluripotent cells.
Publications
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Computer simulation of emerging asymmetry in the mammalian blastocyst. Development (2008) 135(8), 1407-1414
Honda, H., Motosugi, N., Nagai, T., Tanemura, M. and Hiiragi, T.
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Dynamic rearrangement of surface proteins is essential for cytokinesis. Genesis (2008) 46(3), 152-162
Bauer, T., Motosugi, N., Miura, K., Sabe, H. and Hiiragi, T.
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Hypomethylation of parental DNA in the late mouse zygote is not essential for development. Int J Dev Biol (2008) 52(2-3), 295-298
Polanski, Z., Motosugi, N., Tsurumi, C., Hiiragi, T. and Hoffmann, S.
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Stochastic processes during mouse blastocyst patterning. Cells Tissues Organs (2008) 188(1-2), 46-51
Dietrich, J.-E. and Hiiragi, T.
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Bmi1 facilitates primitive endoderm formation by stabilizing Gata6 during early mouse development. Genes & Development (2012) 26(13), 1445–1458
Lavial, F., Bessonnard, S., Ohnishi, Y., Tsumura, A., Chandrashekran, A., Fenwick, Tomaz, R., Hosokawa, H., Nakayama, T., Chambers, I., Hiiragi, T., Chazaud, C., and Azuara, V.
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Gradual meiosis-to-mitosis transition in the early mouse embryo. in Mouse Development – From Oocyte to Stem Cells, in a series of Results and Problems in Cell Differentiation, (2012) 55,107-114
Courtois, A. and Hiiragi, T.
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Stochastic processes in the development of pluripotency in vivo. Biotechnology Journal (2012) 7(6), 737-744
Wennekamp, S. and Hiiragi, T.
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The Kruppel-associated box repressor domain can induce reversible heterochromatization of a mouse locus in vivo. J Biol Chem (2012) 287(30), 25361-25369
Groner, A.C., Tschopp, P., Challet, L., Dietrich, J.-E., Verp, S., Offner, S., Barde, I., Rodriguez, I., Hiiragi, T., and Trono, D.
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The transition from meiotic to mitotic spindle assembly is gradual during early mammalian development. J Cell Biology (2012) 198(3), 357-370
Courtois, A., Schuh, M., Ellenberg, J. and Hiiragi, T.
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A self-organization framework for symmetry breaking in the mammalian embryo. Nat Rev Mol Cell Biol (2013) 14, 454–461
Wennekamp, S., Mesecke, S., Nédélec, F. and Hiiragi, T.
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Cell-to-cell expression variability followed by signal reinforcement progressively segregates early mouse lineages. Nat Cell Biol (2014) 16, 27–37
Ohnishi, Y., Huber, W., Tsumura, A., Kang, M., Xenopoulos, P., Kurimoto, K., Oleś, A. K., Araúzo-Bravo, M. J., Saitou, M., Hadjantonakis, A.-K. and Hiiragi, T.
