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It has long been recognized that mechanical forces underlie mammalian embryonic shape changes. Before gastrulation, the blastocyst embryo undergoes significant shape changes, namely, the blastocyst cavity emerges and expands, and the inner cell mass (ICM) forms and changes in shape. The embryo’s inner pressure has been hypothesized to be the driving mechanical input that causes the expansion of the blastocyst cavity and the shape changes of the ICM. However, how the inner pressure and the mechanics of the trophoblast and the ICM change during development is unknown because of the lack of a suitable tool for quantitative characterization. This work presents a laser-assisted magnetic tweezer technique for measuring the inner pressure and Young’s modulus of the trophoblast and ICM of the blastocyst-stage mouse embryo. The results quantitatively showed that the inner pressure and Young’s modulus of the trophoblast and ICM all increase during progression of mouse blastocysts, providing useful data for understanding how mechanical factors are physiologically integrated with other cues to direct embryo development.

What motivates animal cells to intercalate is a longstanding question that is fundamental to morphogenesis. A basic mode of cell rearrangement involves dynamic multicellular structures called tetrads and rosettes. The contribution of cell-intrinsic and tissue-scale forces to the formation and resolution of these structures remains unclear, especially in vertebrates. Here, we show that Fgfr2 regulates both the formation and resolution of tetrads and rosettes in the mouse embryo, possibly in part by spatially restricting atypical protein kinase C, a negative regulator of non-muscle myosin IIB. We employ micropipette aspiration to show that anisotropic tension is sufficient to rescue the resolution, but not the formation, of tetrads and rosettes in Fgfr2 mutant limb-bud ectoderm. The findings underscore the importance of cell contractility and tissue stress to multicellular vertex formation and resolution, respectively.

The establishment of trophectoderm (TE) manifests as the formation of epithelium, and is dependent on many structural and regulatory components that are commonly found and function in many epithelial tissues. However, the mechanism of TE formation is currently not well understood. Prickle1 (Pk1), a core component of the planar cell polarity (PCP) pathway, is essential for epiblast polarization before gastrulation, yet the roles of Pk family members in early mouse embryogenesis are obscure. Here we found that Pk2−/− embryos died at E3.0–3.5 without forming the blastocyst cavity and not maintained epithelial integrity of TE. These phenotypes were due to loss of the apical–basal (AB) polarity that underlies the asymmetric redistribution of microtubule networks and proper accumulation of AB polarity components on each membrane during compaction. In addition, we found GTP-bound active form of nuclear RhoA was decreased in Pk2−/− embryos during compaction. We further show that the first cell fate decision was disrupted in Pk2−/− embryos. Interestingly, Pk2 localized to the nucleus from the 2-cell to around the 16-cell stage despite its cytoplasmic function previously reported. Inhibiting farnesylation blocked Pk2's nuclear localization and disrupted AB cell polarity, suggesting that Pk2 farnesylation is essential for its nuclear localization and function. The cell polarity phenotype was efficiently rescued by nuclear but not cytoplasmic Pk2, demonstrating the nuclear localization of Pk2 is critical for its function.

Fibroblast Growth Factor (FGF) signaling is known to play an important role during cutaneous development. To elucidate the role of FGF10 during whisker formation, we examined the expression of Fgf10 in normal developing whiskers and phenotypes of Fgf10-deficient whiskers. Fgf10 is first expressed in the maxillary process, lateral and medial nasal processes, then in the mesenchymal cells underneath the future whisker placodes, and in the surrounding mesenchyme of developing whiskers. Fgf10-null whiskers exhibit a significant decrease in number and their structure is disorganized as revealed by scanning electron microscopy. Hair follicle marker genes such as Sonic hedgehog, Patched, and Patched 2 are aberrantly expressed in the mutant whiskers. Thus, FGF10 is required for proper whisker development mediated by SHH signaling in the mouse.

The development of avian cutaneous appendages, feathers and scales, is known to arise from the epithelial–mesenchymal interaction. Here we show that FGF10 is associated with this developmental process as an early signal from mesenchymal cells underlying nascent cutaneous placodes. Expression of Fgf10 was detected in the mesenchymal cells underneath the developing placodes. Forced expression of Fgf10 in the femoral skin suppressed expression of Shh and a zinc finger gene snail-related (cSnR), while induced expression of Bmp2 in the interbud region, resulting in thickening of the epidermal layer. Furthermore, forced expression of Fgf10 in the foot skin caused marked ingrowings of the epidermis. The cells in the epidermal ingrowings expressed β-catenin, proliferating cell nuclear antigen, and an epidermal stem cell marker p63. These results support the idea that FGF10 is a mesenchymally derived stimulator of epidermal development through crosstalk with bone morphogenetic protein (BMP), β-catenin, and other signaling pathways.

In preimplantation mouse development, the first cell lineages to be established are the trophectoderm (TE) and inner cell mass (ICM). The divergence of the two first lineages of the mouse embryo is initiated at the 8-cell stage when blastomeres polarize during compaction. In this process, blastomeres acquire apico-basal polarity typified by apical localization of microvilli and acquisition of cytoplasmic polarity, including asymmetric distribution of E-cadherin and reorganization of the microtubule network. Subsequently, two major interrelated features of TE differentiation required for blastocoel formation include interacellular junction biogenesis and a directed ion transport system, mediated by Na+/K+ ATPase. Previous studies have shown that cell polarity complex (PAR-aPKC complex) regulates the orientation of cell cleavage planes, and cell polarity and adhesion, which altogether can influence the allocation of blastomeres to an outer or inner position in the blastocyst. However, their roles in regulating TE differentiation and blastocyst formation are still unclear. We focus on the roles of Prickle1 and Prickle2, the two mouse homologues of a Drosophila core planar cell polarity gene prickle, during mammalian embryogenesis. We previously reported that the deletion of Prickle1 resulted in early embryonic lethality, between E5.5 and E6.5 with the loss of apico-basal polarity of the epiblast and aberrant dorso-vental patterning of embryo (Tao, H. et al., Proc. Natl. Acad. Sci. USA 106, 2009). On the other hand, we found that Prickle2 knock out mice show that they die at E3.5-4.0 without forming the blastocyst cavity. Detailed analyses revealed that blastomeres after the compaction lost cell polarity that defines inner and outer cells, which gives rise to TE and ICM, respectively. We also found that Prickle2 is preferentially localized to nucleus in early blastomeres. Interestingly, inhibition of fernesylation blocked the nuclear localization and promoted cytoplamic localization of Prickle2, suggesting that the C-terminal fernesylation is essential for tethering Prickle2 to nucleus. Because the cell polarity is disrupted in the inhibitor-treated embryo, nuclear localization of Prickle2 is thought to be prerequisite to the establishment of the polarity. In this meeting, we will discuss about mechanisms that Prickle2 regulates apico-basal polarity during early mouse embryogenesis.