How cells are positioned in embryos: Bringing classical embryology to morphogenesis in C. elegans

Cell rearrangements are crucial during development. We are using C. elegans gastrulation as a simple model to investigate the mechanisms of cell positioning. During C. elegans gastrulation, two endodermal precursor cells (called Ea and Ep, green in figure) move from the ventral surface to the center of the embryo, leaving a gap between these ingressing cells and the eggshell. Six neighboring cells (purple) converge under the endodermal precursors, filling this gap.
 
Using an in vitro system, we found that these movements occurred consistently in the absence of the eggshell and the vitelline envelope. We found that movement of the neighbors toward each other is not dependent on chemotactic signaling between these cells. We further found that C. elegans gastrulation requires intact microfilaments, but not microtubules. The primary mechanism of microfilament-based motility does not appear to be through protrusive structures, such as lamellipodia or filopodia. Instead, our results suggest an alternative mechanism. We found that myosin activity is required for gastrulation, that the apical sides of the ingressing cells contract, and that the ingressing cells determine the direction of movement of their neighboring cells. Based on these results, we have proposed that ingression is driven by an actomyosin-based contraction of the apical side of the ingressing cells, which pulls neighboring cells underneath. We conclude that apical constriction can function to position blastomeres in early embryos, even before anchoring cell-cell junctions form.

Embryonic patterning mechanisms regulate the cytoskeletal machinery that drives morphogenesis, but there are few cases where links between patterning mechanisms and morphogenesis are well understood. We used a combination of genetics, in vivo imaging, and cell manipulations to identify such links in C. elegans gastrulation. We found that ingression of the endodermal precursor cells is regulated by well-studied pathways that specify endodermal cell fate in C. elegans, including a Wnt-Frizzled signaling pathway. We found that Wnt signaling has a role in gastrulation in addition to its earlier roles in regulating endodermal cell fate and cell-cycle timing. In the absence of Wnt signaling, endodermal precursor cells can polarize and enrich myosin II apically, but they still fail to constrict their apical surfaces. We found that a regulatory myosin light chain normally becomes phosphorylated on the apical side of ingressing cells at a conserved site that can lead to myosin-filament formation and contraction of actomyosin networks, and that this phosphorylation depends on Wnt signaling. Based on these results, we concluded that Wnt signaling regulates C. elegans gastrulation through regulatory myosin light-chain phosphorylation, which signals the contraction of the apical surface of ingressing cells.