Description

Results and Discussion

Convergent extension, the mediolateral interdigitation of cells to elongate a tissue array along the anterior-posterior axis, is a conserved feature of embryonic development in animals, and underlies such key events in vertebrates as gastrulation and neurulation. In the Caenorhabditis elegans embryo a convergent extension-like movement known as dorsal intercalation occurs in the embryonic epidermis. The 20 dorsal epidermal cells are born as two rows of 10 cells each that interdigitate to form a single row that straddles the dorsal midline (Williams-Masson et al., 1998). This process, particularly within the posterior dorsal epidermis derived from the C blastomere, is a highly stereotypical process in wild-type embryos (Williams-Masson et al., 1998), making it a promising system for genetic analysis of a simple example of epithelial convergent extension. Dorsal intercalation requires extension of medially directed basolateral protrusions by intercalating cells regulated by Rho family GTPases, including CED-10/Rac, RhoG/MIG-2, and CDC-42, and downstream mediators, which include the WAVE and WASP regulatory complexes (Walck-Shannon et al., 2016; Walck-Shannon et al., 2015).

We sought to identify additional actin regulators that modulate protrusive activity in dorsal cells. A promising candidate is MIG-10, the single MIG-10/RIAM/Lamellipodin (MRL) family member in C. elegans. As the gene name implies, MIG-10 was originally identified in classic genetic screens as a modulator of cell migration (Manser and Wood, 1990). The canonical mutation, mig-10(ct41), results in an amber stop in Exon 3, resulting in predicted severe truncation and strong loss of function of both MIG-10 isoforms (Manser et al., 1997). In neurons, MIG-10 binds activated CED-10/Rac, which polarizes it to the leading edge of growth cones and is necessary for proper axon guidance (Quinn et al., 2008).

Given established requirements for MIG-10 in cell movement and its connection to Rac signaling, we investigated whether MIG-10 plays a significant role during dorsal intercalation in two ways: (1) by endogenously tagging all mig-10 isoforms with mNeonGreen, and (2) by analyzing mig-10 mutants using 4-dimensional DIC microscopy. We endogenously tagged MIG-10 using a C-terminal insertion in the last exon, which is shared by all mig-10 isoforms (Fig. 1A). The endogenous insertion maintains full function of MIG-10, since we did not observe any Mig phenotypes in embryos, larvae, and adults homozygous for the endogenously tagged version of the gene. The earliest prominent expression of MIG-10 we observed was during early morphogenesis in ventral neuroblasts and muscle precursors. The latter organize into four quadrants, two dorsal and two ventral, as embryogenesis proceeds (Viveiros et al., 2011). The two dorsal muscle quadrants lie immediately ventral to the dorsal epidermis, and show prominent expression of MIG-10::mNG (Fig. 1B, 1C). While it is possible that there is low-level expression of mig-10 in dorsal epidermal cells, if it is expressed there it is present at much lower levels than in dorsal muscle quadrants. We did not see any overlap between epidermally driven LifeAct::mCherry and MIG-10::mNeonGreen (Fig. 1E), further suggesting that detectable dorsal mig-10 expression is restricted to muscle precursors at this stage of development.

The relative lack of expression of MIG-10 in the dorsal epidermis suggests that it is not a major contributor to actin dynamics during dorsal intercalation. We confirmed this using 4d DIC microscopy. mig-10(ct41) homozygotes complete dorsal intercalation in a manner indistinguishable from wild-type embryos (Fig. 1D). In 8 mounts with 21 dorsal presenting embryos, we observed no dorsal intercalation defects in the mig-10(ct41) background, a result indistinguishable from wild-type embryos (no defects in 7 mounts with 26 dorsal presenting wild-type embryos).

Taken together, these results suggest that MIG-10 is not an essential actin regulator in dorsal epidermal cells during dorsal intercalation. It will be interesting to determine if MIG-10 acts redundantly with other actin regulators or if it plays more prominent roles during other cell migration events in the embryo.