All authors wrote the manuscript. Acknowledgments We thank Benjamin Rishirumuhirwa and Alen Tosenberger for fruitful discussions. G.D. simulations of a human population of 25 cells under numerous experimental conditions to compare their outcome with that of mutant embryos or of embryos submitted to exogenous treatments that GANT61 interfere with Fgf signaling. The results are analyzed by means of bifurcation diagrams. Finally, the model predicts that heterogeneities in extracellular Fgf4 concentration play a primary part in the spatial set up of the Epi/PrE cells inside a salt-and-pepper pattern. If, instead of heterogeneities in extracellular Fgf4 concentration, internal fluctuations in the levels of manifestation of the transcription factors are considered as a source of randomness, simulations forecast the event of unrealistic switches between the Epi and the PrE cell fates, as well as the development of some cells toward one of these claims without moving through the previous ICM state, in contrast to what is definitely observed in?vivo. Intro During early murine embryogenesis, two differentiation processes take place before the implantation of the egg in the uterus. The 1st one gives rise to the inner cell mass (ICM) and the trophectoderm (TE), which communicate Oct4 and Cdx2, respectively. The second differentiation process corresponds to the specification of ICM cells into primitive endoderm (PrE) and epiblast (Epi) cells. Whereas PrE and TE cells contribute to the formation of extraembryonic cells, such as the placenta, Epi cells primarily give rise to the embryo itself. The epiblast is also the cellular compartment from which embryonic stem (Sera) cells can be derived. Sera cells are priceless tools in a wide range of medical applications. For these reasons, understanding the molecular mechanisms leading to the formation of Epi cells constitutes an important goal in developmental biology. The differentiation of ICM cells into Epi and PrE is definitely controlled by two antagonistic transcription factors, Nanog and Gata6. Nanog is necessary to produce Epi cells (1, 2, 3, 4), whereas Gata6 is required for the specification GANT61 of PrE cells both in?vitro and in?vivo (5, 6). These genes start to become zygotically expressed round the two/four-cell stage (related to embryonic day time E1.5CE2), and both proteins can be detected in most ICM cells from the eight-cell stage (E2.5). Between the 8- and 32-cell phases, Nanog and Gata6 proteins are coexpressed at increasing levels in almost all ICM cells (7, 8, 9, 10). Then, from E3.0CE3.25, their expression patterns start to become mutually exclusive. As a consequence, at E3.75, two distinct cell typesdistributed inside a salt-and-pepper patternconstitute the ICM: Gata6-expressing PrE progenitors and Nanog-expressing Epi progenitors (8, 9, 11, 12, 13). Later on, these two populations are sorted out so that PrE cells form an epithelium that separates the Epi cells from your blastocoel (8, 14, 15, 16). The Fgf/Erk signaling pathway offers been shown to bias the Epi/PrE fate choice during embryonic development. Indeed, the proper specification of PrE requires the expression of the Fgf receptor the Fgf ligand Fgf4, and the Erk adaptor (12, 17, 18, 19, 20, 21, 22). Moreover, between E3.0CE3.25 and E4.0, ICM cells can GANT61 be forced to differentiate into a specific fate (Epi or PrE) in response to exogenously induced, nonphysiological variations in Fgf/Erk signaling. Indeed, culturing wild-type (WT) embryos with inhibitors of the Fgf/Erk signaling pathway prospects to the absence of?PrE cells, whereas culturing them with recombinant Fgf4 induces a severe reduction in the number of Epi cells (23, 24). To support the experimental investigation of the complex tasks of Nanog, Gata6, and Fgf signaling in determining the Epi or PrE cell fates, we previously built a computational model describing the gene regulatory network (GRN) controlling ICM specification (6). We found that in appropriate conditions, this model exhibits three stable stable claims (tristability), which correspond to ICM, Epi, and PrE cells, respectively. Computational simulations replicated or expected a variety of cell behaviors observed in different experimental conditions, namely, 1) the self-organized development of a human population of 25 cells GANT61 toward the ICM-like state 1st, followed by a specification into Epi or PrE cells (reaching similar amounts and showing a salt-and-pepper pattern); 2) required differentiation into a specific fate (Epi or PrE) in response to the exogenously induced variations in Fgf/Erk signaling explained in the previous paragraph; 3) specification into the Epi phenotype for those cells in the Gata6?/? mutant, as well as a partial deficit in the specification of Rabbit Polyclonal to POLR1C PrE cells in Gata6+/? embryos; and 4) faster specification into Epi cells in Gata6.