This work has focused on improving our knowledge of the thyroid specification process. Thyroid cells are derived from mouse embryonic stem cells (mESCs) by directed differentiation through multiple intermediate developmental stages, including anterior foregut endoderm (AFE), prior to NKX2-1+ thyroid progenitor specification. To investigate if transient Nkx2-1 expression can increase the efficiency of thyroid specification, we utilized a mESC line double knock-in GFP-T/hCD4-Foxa2 with a doxycycline inducible (Tet-On) Nkx2-1 transgene. Transient activation of the Nkx2-1 transgene at the AFE stage leads to stable induction of high levels of endogenous Nkx2-1 as well as early and mature thyroid-specific markers including Pax8, Foxe1, Tg, Nis, and Tshr. Lung and neuronal NKX2-1+ lineages were not derived in this system. The thyroid progeny mature and organize into follicle-like structures in 3D culture. These follicles express adherens and tight junction proteins indicative of an epithelial character and produce the thyroid hormone thyroxine (T4) in the presence of iodide. Critical determinants of thyroid lineage specification have been revealed by variations in developmental stage timing, signaling pathways, and sorting of AFE-stage subpopulations. To provide further insights into the mechanisms of thyroid specification, RNA-Seq data acquired from relevant stages has identified potential genes involved in early thyroid development. The results demonstrate that Nkx2-1 can act as a stage-specific inductive signal during directed differentiation of mESCs to thyroid follicular cells. We have also developed a mouse model to recapitulate these results in an in vivo context. This work has provided novel insights into thyroid specification and provides an efficient system for deriving and studying thyroid cells, which can be used for in vitro modeling of development and disease.

A E3.5 jours de développement (E3.5), l’embryon murin se compose d’une monocouche de cellules externes correspondant au Trophectoderme (TE) et d’une masse cellulaire interne (MCI), hétérogène, constituée de deux sous-populations de cellules précurseurs : les cellules épiblastiques (Epi) et les cellules d’endoderme primitif (EPr). NANOG, marqueur épiblastique et GATA6, marqueur de l’EPr, sont co-exprimés à E3.5 dans la MCI puis adoptent une expression exclusive au sein de leur lignage respectif. La différenciation du lignage EPr nécessite l’expression de GATA6 et l’activation de la voie Récepteur Tyrosine Kinase (RTK) activée par le FGF (RTK-FGF) pour l’induction de gènes cibles de GATA6 tels que Sox17 et Gata4.Au cours de ma thèse, j’ai, dans un premier temps, étudié la relation GATA6/voie RTK-FGF lors de l’induction de l’expression des gènes de différenciation de l’EPr. J’ai utilisé des cellules souches embryonnaires murines ES sauvages ou mutantes pour Gata6 (ES Gata6-/-), dans lesquelles j’ai surexprimé différentes formes mutantes de Gata6 inactivées sur les différents résidus identifiés comme potentiellement phosphorylables par la voie RTK-FGF. Ainsi, j’ai analysé l’expression protéique des gènes Sox17 et Gata4 ainsi que des expressions ARN de ces cibles et d’autres gènes caractéristiques exprimés dans l’EPr dans les différentes conditions de surexpression des formes de Gata6 en absence ou présence d’inhibiteurs de la voie RTK-FGF. Ainsi, j’ai pu mettre en évidence que la transmission du signal s’effectue au travers de récepteur au FGF et qu’il existe une compensation entre les branches RTK-MEK-ERK et RTK-PI3K ciblant le résidu Sérine 37 de GATA6. Enfin, les résidus S34 et T509 sont nécessaires et les résidus S34, S37 et T509 semblent coopérer, au travers d’un mécanisme pour le moment non détaillé, pour l’induction des gènes cibles exprimés au sein de l’EPr.Dans un second temps, j’ai débuté la caractérisation phénotypique du rôle des facteurs Dickkopf1 (DKK1), un inhibiteur de la voie WNT/β-caténine, et NOGGIN, un inhibiteur de la voie des Bone Morphogenic Protein (BMP) lors de la différentiation de l’EPr en endoderme pariétal (EP) et viscéral (EV). A l’aide de modèles de souris KO pour DKK1 et NOGGIN, croisées en fond C57Bl6 pur, j’ai pu observer que l’expression d’OCT4 était maintenue au sein des embryons homozygotes mutants pour Dkk1 et double homozygotes mutants pour Dkk1 et Noggin. Cependant, le mécanisme potentiel de compensation ou de coopération de ces deux marqueurs n’est pour le moment pas détaillé précisément et mérite l’analyse d’un plus grand nombre d’embryons mutants. / At E3.5 days of development (E3.5), mouse embryo consists of a monolayer of external cells corresponding to Trophectoderme (TE) and of an intern cell mass (ICM), heterogeneous, constituted by two subpopulations of precursory cells: epiblastic cells (Epi) and primitive endoderm cells (EPr). NANOG, an Epi marker and GATA6, a PrE marker, are co-expressed at E3,5 in the MCI and then adopt an exclusive expression within their respective lineage. EPr differentiation requires both expression of GATA6 and RTK pathway, activated by FGF ligand, in order to induce late markers Sox17 and Gata4 expression.First, I studied the relation GATA6/RTK during this process to understand the mechanism of induction of these target genes during final EPr differentiation. I used embryonic stem cells ES WT or Gata6 mutants (ES Gata6-/-), in which I transfected various Gata6 mutant constructions on different residues characterized as potentially phosphorylable by the RTK pathway. So, I analyzed protein expression of Sox17 and Gata4 target genes as well as RNA expression of characteristic genes expressed in the EPr in different inhibition conditions of RTK pathway. So, I was able to highlight that the transmission of the signal is made through the FGF receptor (FGFR1) and that there is compensation between RTK-MEK-ERK and RTK-PI3K pathways highlighted by later Gata6 overexpression of certain mutant forms. Finally, residue S34, S37 and T509 seems to cooperate, through a mechanism not detailed for the moment, for the induction of the EPr target genes.Then, I was interested to phenotypically characterize the role of Dickkopf1 (DKK1), an inhibitor of the WNT/β-catenin pathway, and NOGGIN, an inhibitor of the Bone Morphogenic Protein (BMP) pathway during the EPr differentiation in parietal endoderm (EP) and visceral (EV). Using models of mouse KO for Dkk1 and Noggin, met in pure background C57Bl6, I was able to observe that OCT4 expression was maintained within the Dkk1-/-, and Dkk1-/- Noggin-/- embryos. However, the potential compensation or cooperation mechanism of these two markers is not understanding well for the moment and deserves the analysis of a largest mutant embryos number.

A 3,5 jours de développement (E3.5), l'embryon de souris est composé d'une couche externe de trophectoderme (TE) entourant la cavité blastocélique et la masse cellulaire interne (MCI). La MCI comprend une population hétérogène de précurseurs d'épiblaste (Epi) et d'endoderme primitif (EPr) dans une configuration "poivre et sel". A E4.5, ces cellules ségrégent, les cellules d'EPr migrant vers la surface de la MCI pour former un épithélium. A E4.75 cet épithélium donne naissance à 2 tissus distincts : un épithélium d'endoderme viscéral (EP) et à l'endoderme pariétal (EP) qui migre le long du TE. Une transition épithélium-mésenchyme est impliquée dans la formation de l'EP. Je m'intéresse au rôle de Dkk1, un inhibiteur de la voie Wnt canonique et activateur de la voie Wnt/PCP, et Noggin, un inhibiteur des BMP, dans la différenciation de l'endoderme extraembryonnaire. J'ai montré que Dkk1 est un marqueur de l'EPr qui devient apicalement polarisé à E4.5. Son expression est ensuite restreinte aux cellules de l'EP. J'ai aussi montré que Noggin est exprimé dès la préimplantation puis dans l'EP et au niveau de la charnière EV-Ep. Par des expériences de perte et de gain de fonction des voies Wnt et BMP et en utilisant les souris mutantes j'ai analysé le rôle de ces deux facteurs dans la différenciation de l'endoderme extraembryonnaire. / At 3.5 days of development (E3.5), the mouse embryo consists of an outer layer of trophectoderm (TE) surrounding the blastocelic cavity and the inner cell mass (ICM). The ICM is composed of intermingled populations of epiblast (Epi) and primitive endoderm (PrE) precursors, that sort to form two distinct tissues. At E4.75 this epithelium differentiates into visceral endoderm (VE) and parietal endoderm (PE) that migrates along TE. An epithelium-mesenchyme transition (EMT) is involved in PE formation while the VE is maintained as an epithelium. My work focuses on the role of Dkk1, a Wnt canonical pathway inhibitor and Wnt/PCP pathway activator, and Noggin, a BMP pathway inhibitor, in extraembryonic endoderm differentiation. I have shown that Dkk1 is a marker of PrE precursors and is apically polarised at E4.5. Afterwards, its expression is restricted to PE. Noggin is expressed during preimplantation and then in PE and EV-EP hinge. By gain and loss of fonction experiments of Wnt and BMP pathways and by using mutant mice, I studied the role of these two factors in extraembryonic endoderm differentiation.

<p>The present study was conducted in order to assess a role of the SH2 domain-containing adapter protein SHB in development and cell differentiation.</p><p>Embryonic stem (ES) cells overexpressing SHB and SHB with an inactive SH2 domain (R522K-SHB) were obtained. Microarray analysis in the SHB clone revealed altered expression of genes connected with neural cell function. The R522K-SHB clone exhibited altered expression of several transcription factors related to development. ES cells were differentiated by forming aggregates named embryoid bodies (EBs). The morphology of EBs was altered in the R522K-SHB clones, which showed fewer cavities. Expression of endodermal markers was decreased in the R522K-SHB EBs. </p><p>To further investigate the role of SHB in differentiation, murine ES cell lines deficient for one (SHB+/-) or both SHB alleles (SHB-/-) were generated. SHB deficient clones increased the expression of mesendodermal and endodermal markers and decreased expression of two receptors, VEGFR2 and FGFR1, connected with blood vessel differentiation. Similarly, blood vessels showed an altered morphology in SHB+/- and SHB-/- EBs after VEGF stimulation. SHB-/- ES cells also formed fewer blood colonies than control ES cells.</p><p>Finally, the role of the SHB adapter protein in vivo was analyzed by generating a SHB deficient mouse (SHB-/-). SHB-/- animals are viable, fertile, but suffer from leukopenia and anemia. SHB-/- animals demonstrate an abnormal morphology of blood vessels in the liver and kidney. Breeding of SHB+/- animals revealed an abnormal segregation of the mutant allele with an increased number of SHB+/- animals and a decreased number of SHB-/- and SHB+/+animals. Backcross analysis of SHB+/- females with SHB+/+ males displayed an increased number of SHB+/- offspring already at the blastocyst level. Simultaneously, embryos from SHB+/- mothers show an increased malformation rate in comparison to embryos from SHB+/+ mothers.</p><p>In summary, the study suggests a role of SHB in reproduction and development and in mesodermal and endodermal specification. </p>

The present study was conducted in order to assess a role of the SH2 domain-containing adapter protein SHB in development and cell differentiation. Embryonic stem (ES) cells overexpressing SHB and SHB with an inactive SH2 domain (R522K-SHB) were obtained. Microarray analysis in the SHB clone revealed altered expression of genes connected with neural cell function. The R522K-SHB clone exhibited altered expression of several transcription factors related to development. ES cells were differentiated by forming aggregates named embryoid bodies (EBs). The morphology of EBs was altered in the R522K-SHB clones, which showed fewer cavities. Expression of endodermal markers was decreased in the R522K-SHB EBs. To further investigate the role of SHB in differentiation, murine ES cell lines deficient for one (SHB+/-) or both SHB alleles (SHB-/-) were generated. SHB deficient clones increased the expression of mesendodermal and endodermal markers and decreased expression of two receptors, VEGFR2 and FGFR1, connected with blood vessel differentiation. Similarly, blood vessels showed an altered morphology in SHB+/- and SHB-/- EBs after VEGF stimulation. SHB-/- ES cells also formed fewer blood colonies than control ES cells. Finally, the role of the SHB adapter protein in vivo was analyzed by generating a SHB deficient mouse (SHB-/-). SHB-/- animals are viable, fertile, but suffer from leukopenia and anemia. SHB-/- animals demonstrate an abnormal morphology of blood vessels in the liver and kidney. Breeding of SHB+/- animals revealed an abnormal segregation of the mutant allele with an increased number of SHB+/- animals and a decreased number of SHB-/- and SHB+/+animals. Backcross analysis of SHB+/- females with SHB+/+ males displayed an increased number of SHB+/- offspring already at the blastocyst level. Simultaneously, embryos from SHB+/- mothers show an increased malformation rate in comparison to embryos from SHB+/+ mothers. In summary, the study suggests a role of SHB in reproduction and development and in mesodermal and endodermal specification.

Shb is a ubiquitously expressed adaptor protein with the ability to bind several tyrosine kinase receptors and intracellular signaling proteins. Previous studies have implied a wide spectrum of Shb-mediated cellular responses, which motivated me to further investigate the role of Shb in differentiation and angiogenesis. Embryonic stem (ES) cells differentiate into endoderm and mesoderm from a bipotent mesendodermal cell population. Interregulatory signals between these germlayers are required for further specification. ES cells overexpressing Shb with an inactive SH2 domain (R522K-Shb) altered the expression of endodermal genes as a consequence of upregulated FGF expression. This response was enhanced by addition of activin A, suggesting a synergistic mechanism operative between FGF and activin A signaling in endoderm specification. To investigate a role for Shb in mesodermal specification, Shb knockout ES cells were established. These cells showed a reduced ability to form blood vessels after VEGF stimulation and delayed downregulation of genes associated with mesendoderm, indicating a reduced capacity for these cells to enter later stages. To assess a role for Shb in tumor cell apoptosis, Shb expression was silenced in angiosarcoma endothelial cells. FAK-phosphorylation was reduced in Shb knockdown cells and this made them more susceptible to apoptotic stimuli both in vitro and in vivo. Shb knockout microvasculature in mouse kidney, liver, and heart showed irregular endothelial linings with cytoplasmic projections toward the lumen, a feature that was also related to increased vascular permeability. VEGF treatment failed to stimulate vascular permeability in Shb knockout mice. In order to elucidate whether these features relate to reduced angiogenesis, tumor growth was examined. Tumors grown in knockout mice showed reduced growth capacity and lower vessel density. In conclusion, Shb is a multifunctional adaptor protein that may be involved in several cellular responses both during embryonic development and adult life.

The Drosophila melanogaster gene Anaplastic Lymphoma Kinase (DAlk) is homologous to mammalian Alk, which is a member of the Alk/Ltk family of receptor tyrosine kinases (RTKs). In humans the t(2;5) translocation involving the Alk locus encodes an active form of Alk that is the causative agent in Non-Hodgkin’s Lymphoma (Morris et al., 1994). Alk has also been associated with other cancers such as inflammatory myofibroblastic tumours (IMTs). The physiological function of the Alk RTK has not been described in any system until very recently, and is still not defined in vertebrates. The molecular similarity between Drosophila Alk and mammalian Alk suggested that mutation of Alk in flies may affect similar functional and developmental processes, and thus lead to some understanding of Alk function in vivo. By employing an EMS mutagenesis screen we were able to obtain loss-of-function mutants in the Drosophila DAlk gene. Eleven independent DAlk mutants were identified and characterized. DAlk is normally expressed in the developing gut and in the CNS. DAlk mutant animals have a lethal phenotype and die at late embryonic stages or as 1st instar larva. In DAlk mutant embryos there is a complete failure in the development of the midgut whereas the CNS appears normal. The midgut consists of visceral musculature that is syncytial and is formed by fusion of multiple myoblasts. This is a dynamic process where two types of myoblasts, i.e. fusion-competent-myoblasts and founder-cells that function as seeds for muscle formation, fuse. In DAlk homozygous embryos there is no founder cell specification, which explains the failure of midgut formation in these embryos. Recently a novel secreted molecule Jelly Belly (Jeb) was identified. Jeb is expressed in the tissue neighbouring the DAlk expressing cells of the developing visceral mesoderm. Jeb mutant embryos show a phenotype that is similar to that of DAlk mutant embryos. We have been able to show that Jeb is the ligand for DAlk in the developing visceral mesoderm and that Jeb binding stimulates a DAlk driven ERK signaling pathway. This leads to the expression of Dumbfounded (duf)/kin of Irregular chiasm-C (kirre), a founder-cell specific immunoglobulin that has an important role in myoblast aggregation and fusion. The functional Drosophila midgut is made up of the visceral muscle that encircles the endodermal tube. This tube formation includes migration of cells originating in the anterior and posterior parts of the embryo, first along the anterior-posterior axis using the visceral mesoderm as a template, then dorsally and ventrally. In DAlk mutant embryos there is no visceral muscle fusion and both the visceral mesoderm and the endoderm fail to undergo dorsal-ventral migration.

One of the fundamental questions in developmental biology is how cells communicate during embryonic development to pattern the animal with defined axes and correctly placed organs. There are several key signal transduction pathways whose signaling has been found to be crucial during this period in the life history of many model organisms and whose functions have been well conserved between species. Two of those are the Notch and Hedgehog signal transduction pathways. Previous work established that the Notch pathway is important in the specification of mesoderm in the sea urchin embryo. Here it is established that the Hedgehog pathway is important for mesoderm patterning in the echinoderm embryo.In many animals, including the sea urchin, endomesoderm is specified as a bipotential tissue which is then subdivided through cell signaling to become endoderm and mesoderm. Notch signaling was found to be critical for that dichotomy; endomesoderm that received the Notch signal becomes mesoderm, the remaining endomesoderm becomes endoderm. Prior to this work, no functional roles for Hedgehog signaling in the sea urchin had been defined, though this pathway is known to operate in organisms throughout the animal kingdom. Here we find through analysis and comparison of the sea urchin genome with cnidarians, arthropods, urochordates, and vertebrates that key components and modifiers of the Notch and Hedgehog signaling pathways are well conserved among metazoans. Many animals contain the full suite of genes that constitute both pathways, and in deuterostomes the pathways operate in embryos to mediate similar fate decisions. The Notch pathway, for example, is engaged in endomesoderm gene regulatory networks and in neural functions. In the sea urchin RNA in situ hybridization of Notch pathway members confirms that Notch functions sequentially in the vegetal-most secondary mesenchyme cells and later in the endoderm.The Hh signaling pathway is essential for patterning of many structures in vertebrates ranging from the nervous system, chordamesoderm, and limb to endodermal organs. In the sea urchin, a basal deuterostome, we show that Hedgehog (Hh) signaling participates in organizing the mesoderm. During gastrulation expression of the Hh ligand is localized to the endoderm while the co-receptors Patched (Ptc) and Smoothened (Smo) are expressed in the neighboring secondary mesoderm and in the ventrolaterally clustered primary mesenchyme cells where skeletogenesis initiates. Perturbations of Hh signaling cause embryos to develop with skeletal defects, as well as inappropriate secondary mesoderm patterning, although initial specification of secondary mesoderm occurs normally. Perturbations of Hedgehog pathway members altered normal numbers of pigment and blastocoelar cells, randomized left-right signaling in coelomic pouches, and resulted in disorganization of the circumesophageal muscle, causing an inability to perform peristaltic movements. Together our data support the requirement of Hh signaling in patterning each of the mesoderm subtypes in the sea urchin embryo.Activation of the Hedgehog pathway requires FoxA acting upstream of Hedgehog transcription, early in gastrulation. When FoxA is knocked-down there is a loss of transcription of Hedgehog and Hh expression is expanded in embryos expressing ectopic FoxA. In collaboration with another lab, we found that FoxA acts to repress mesodermal genes within the endoderm as part of the endomesoderm dichotomy. If FoxA expression is reduced by a morpholino, more endomesoderm cells become pigment and other mesenchymal cell types, and less gut is specified. Conversely, when FoxA is ectopically expressed, endoderm is increased at the expense of mesoderm. More specifically we found through mosaic analysis that FoxA acts in a portion of the endomesoderm derived from one of two tiers of vegetal cells at the 60 cell stage called the veg2 cells. FoxA remains on in all endoderm and its territory of expression is superimposeable with the location of Hh expression.The data we present here together with previous studies suggest a model in which Notch signaling cues cells of the endomesoderm to become mesoderm, while cells of the nascent endoderm upregulate FoxA. FoxA ensures proper partitioning of endoderm from mesoderm by repressing mesoderm genes, as well as positively regulating transcription of Hedgehog in the endoderm. The Ptc and Smo transducing apparatus is separately expressed in mesoderm. Hh then signals to its receptors in the mesoderm to convey patterning information of tissues derived from that mesoderm. Thus, Hh, Ptc and Smo molecules diverge during specification then converge during signaling to play important roles in mesoderm development in the sea urchin. / Dissertation

The fully developed and functional Drosophila midgut comprises two layers, the visceral mesoderm and the endoderm. The visceral muscle of the midgut is formed by the fusion of founder cells with fusion competent cells to form the muscle syncytia. The specification of these cells and thus the fusion and the formation of the midgut muscle is dependent on the  Receptor tyrosine kinase (RTK) Alk (Loren et al., 2003). The endoderm underlies the visceral muscle and is formed from cells that originate from the anterior and the posterior parts of the embryo. These cells use the visceral mesoderm as a substrate for their migration. Using Alk mutant animals, we have studied endoderm migration during embryonic development. While the initial migration of the endoderm is not affected in the absence of the visceral mesoderm, we observe that the later dorsal-ventral endodermal migration does not take place. The development of the visceral muscle and its dependence on the endoderm is poorly understood.  We have analysed gürtelchen (gurt) mutant animals, originally identified in a genetic screen for mutations affecting visceral muscle formation. Gurt mutants are so named due to their belt-like phenotype of the visceral muscle (gürtelchen is German for belt). Mapping of the genomic locus identified gurt as a mutation in a previously described gene - huckebein (hkb) which is known to have an important function in endoderm development. Gurt (hkb) mutants were used to further study the interaction between the endoderm and the visceral muscle during development. The initial specification of founder cells and fusion competent myoblasts as well as fusion events are unaffected in gurt (hkb) mutants, however, the elongation and stretching of the visceral muscle does not proceed as normal. Moreover, ablation of the visceral mesoderm disrupts endoderm migration, while ablation of the endoderm results in a delayed disruption of visceral muscle formation. Signaling between the two tissues was investigated in detail. Since Alk is a critical player in visceral muscle development, we employed Alk mutant embryos for this task. In addition to the role of Alk in specifying the founder cells and initiating the visceral muscle fusion, we have shown that Alk mediated signaling has a role in the induction of the midgut constriction process by regulating dpp expression in the developing embryonic gut.  Finally, we wished to identify genes in the founder cells/fusion competent myoblasts that might be regulated by Alk. C3G is a gunaine nucleotide exchange factor expressed in the visceral muscle founder cells. Deletion of the Drosophila C3G locus resulted in the generation of null mutants in C3G which are viable, but display decreased longevity, fitness and are semi-lethal. Further analysis of C3G mutants indicated that C3G is essential for normal larval musculature development, in part by regulating integrin localization at muscle attachment sites.

The preimplantation stages of mammalian development are dedicated to the differentiation of two extraembryonic epithelia, the trophectoderm (TE) and the primitive endoderm (PrE), and their segregation from the pluripotent embryonic lineage, the epiblast. The TE and PrE are responsible for implantation into the uterus and for producing the tissues that will support and pattern the epiblast as it develops into the foetus. PrE and epiblast are formed in a two step process that involves random cell fate specification, mediated by fibroblast growth factor (FGF) signalling, and cell sorting through several mechanisms. In the present work I have addressed aspects of both steps of this process. Chimaera assays showed that epiblast precursors transplanted onto a recipient embryo rarely differentiate into PrE, while PrE precursors are able to switch their identity and become epiblast. Transient stimulation or inhibition of the FGF4-ERK pathway in the chimaeras can modify the behaviour of these cells and restore the plasticity of epiblast precursors. This work shows that epiblast precursors are refractory to differentiation signals, thus ensuring the preservation of the embryonic lineage. I have also found that atypical Protein Kinase C (aPKC) is a marker of PrE cells and that pharmacological inhibition of aPKC impairs the segregation of PrE and epiblast precursors. Furthermore, it affects the survival of PrE cells and can alter the subcellular localisation of the PrE transcription factor GATA4. These data indicate aPKC plays a central role for the sorting of the PrE and epiblast populations and links cell position within the embryo to PrE maturation and survival. Lastly, I have found that aPKC can directly phosphorylate GATA4 in vitro. Knockdown of GATA4 affects cell position within the embryo, whereas aPKC knockdown reduces the number of GATA4-positive cells. These results indicate GATA4 plays an important role in cell sorting during preimplantation development and suggest phosphorylation by aPKC could determine its presence in the nuclei of PrE cells. My work, in the light of the current knowledge, supports a model where the earliest cell fate decisions during mammalian development depend on cellular interactions and not on inherited cell fate determinants. This robust mode of development underlies the plasticity of the preimplantation embryo and ensures the formation of the first mammalian cell lineages, critical for any further progression in mammalian development.