Figure 8

Rho/Rock signaling stabilize membrane-bound N-cadherin while inhibiting neural crest (NC) delamination. (A) In control explants, N-cadherin (red) is expressed in the membrane of epithelial and epithelioid cells flattening onto the substrate but is downregulated upon delamination in mesenchymal cells. (B) N-cadherin disappears from the membrane of Y27632-treated cells that exhibit a mesenchymal phenotype even close to the neural tube (NT). (C) Lysophosphatidic acid (LPA) strenghtens membranous N-cadherin immunoreactivity while inhibiting epithelial-to-mesenchymal transition (EMT). (D) These effects of LPA are reverted by co-treatment with Y27632. (E,F) Inhibition of N-cadherin cleavage by GI254023X keeps membrane-associated N-cadherin and prevents NC EMT; both effects are reverted by inhibiting Rock activity. Note in (F) that membrane N-cadherin staining is either fragmentary or absent. Insets show low magnification of phase contrast images. In all panels, the NT explant is to the top. (G,H) Treatment with LPA/pluronic gel in vivo prevents the normal downregulation of N-cadherin from adherens junctions (arrowhead in (H)) that is seen in control gel-treated embryos (arrows in (G)) at dissociated levels of the axis. (I and inset) Loss of RhoB function by transfection of N19-RhoB/green fluorescent protein (GFP) prematurely downregulates N-cadherin from adherens junctions in the apical NT when compared to the untreated side (arrowhead pointing at the apical side of the NT expressing N19/GFP but devoid of N-cadherin). Note that electroporation at more ventral domains of the NT was without effect on N-cadherin staining (ventral to the arrow), consistent with the dorsally restricted expression of RhoB. DM, dermomyotome; Scl, sclerotome. Bar: 15 μM (A-F); 500 μM (insets in E,F); 26 μM (G,H); 14 μM (I); 65 μM, inset in (I).