HIPPO Pathway Members Restrict SOX2 to the Inner Cell Mass Where It Promotes ICM Fates in the Mouse Blastocyst

Abstract

Pluripotent epiblast (EPI) cells, present in the inner cell mass (ICM) of the mouse blastocyst, are progenitors of both embryonic stem (ES) cells and the fetus. Discovering how pluripotency genes regulate cell fate decisions in the blastocyst provides a valuable way to understand how pluripotency is normally established. EPI cells are specified by two consecutive cell fate decisions. The first decision segregates ICM from trophectoderm (TE), an extraembryonic cell type. The second decision subdivides ICM into EPI and primitive endoderm (PE), another extraembryonic cell type. Here, we investigate the roles and regulation of the pluripotency gene Sox2 during blastocyst formation. First, we investigate the regulation of Sox2 patterning and show that SOX2 is restricted to ICM progenitors prior to blastocyst formation by members of the HIPPO pathway, independent of CDX2, the TE transcription factor that restricts Oct4 and Nanog to the ICM. Second, we investigate the requirement for Sox2 in cell fate specification during blastocyst formation. We show that neither maternal (M) nor zygotic (Z) Sox2 is required for blastocyst formation, nor for initial expression of the pluripotency genes Oct4 or Nanog in the ICM. Rather, Z Sox2 initially promotes development of the primitive endoderm (PE) non cell-autonomously via FGF4, and then later maintains expression of pluripotency genes in the ICM. The significance of these observations is that 1) ICM and TE genes are spatially patterned in parallel prior to blastocyst formation and 2) both the roles and regulation of Sox2 in the blastocyst are unique compared to other pluripotency factors such as Oct4 or Nanog.

Lineage specification in the mouse preimplantation embryo

“…The suppression of SOX2 expression in the TE is CDX2 independent and thus differs from NANOG and OCT4 regulation . This indicates that YAP activates a distinct repression mechanism in outer cells for the regulation of SOX2 expression (Wicklow et al, 2014).…”

“…Interestingly, the YAP/Hippo signalling cascade regulates not only TE-specific genes but also the ICM-restricted expression of SOX2, which is one of earliest markers of inner cells. Indeed, preventing YAP nuclear localization in outer TE cells is sufficient to induce ectopic SOX2 expression (Wicklow et al, 2014). The suppression of SOX2 expression in the TE is CDX2 independent and thus differs from NANOG and OCT4 regulation .…”

“…Another EPI-specific marker, SOX2, is also necessary for PE maturation but is not required for EPI/PE specification. Indeed, Sox2 mutant embryos are able to generate NANOG-expressing and GATA6-expressing cells in the correct proportions; however, the level of GATA6 expression is lower and there are fewer cells expressing SOX17 at E3.75 (Wicklow et al, 2014). By E4.25, the number of SOX17-expressing cells is restored, suggesting that the expression of SOX17 is simply delayed in PE cells in these mutants.…”

Cheetah interspecific SCNT followed by embryo aggregation improves in vitro development but not pluripotent gene expression

“…In the mouse, it was demonstrated that Oct4 in association with Sox2 and Nanog forms a complex, which maintains the pluripotent cells in the ICM of the embryos (Nichols et al 1998, Mitsui et al 2003, Rodda et al 2005, Masui et al 2007. Moreover, Sox2 expression is necessary during embryogenesis to facilitate establishment of the yolk sac lineage, which is essential for gestation (Wicklow et al 2014). In the mouse, the differentiation of the ICM and the trophectoderm (TE) is also directed by the antagonistic expression of Oct4 and Cdx2.…”

BRG1 Governs Nanog Transcription in Early Mouse Embryos and Embryonic Stem Cells via Antagonism of Histone H3 Lysine 9/14 Acetylation

“…During blastocyst formation, the expression of key transcription factors, such as octamer-binding transcription factor 4 (OCT4), Nanog homeobox (NANOG), and sex-determining region Y box 2 (SOX2), becomes restricted to the pluripotent ICM, while transcription factor AP-2 gamma (TFAP2C), GATA binding protein 3 (GATA3), and caudal type homeobox 2 (CDX2) are expressed exclusively in the trophoblast lineage (4)(5)(6)(7)(8)(9)(10). The spatial and temporal expression of these lineage-specific factors is controlled by position-dependent HIPPO signaling, transcription factor regulatory loops, and chromatin modifications (2,3,(10)(11)(12). For example, the HIPPO signaling pathway differentially regulates lineage formation via the downregulation of CDX2 expression in the ICM and SOX2 expression in the trophoblast (10,12).…”