Relationship between apical contractions and extension advancement. (A)(Left) Spatiotemporal correlation of contractions and extension advancement in a representative embryo. Contractions are outlined with a dashed white line, while initial and final position of neighboring cell extensions are labeled with a black and red dashed line, respectively. Red arrows indicate the direction of extension advancement. (Right) Schematic representation of multiple contractions in a representative embryo. Colored areas represent initial spread of contractile foci that cluster into one focus, and arrows indicate contractions coupled to extension advancement; the order of contractions is color-coded. (B) Graphs correlating integrated local fluorescence intensity of non-muscle myosin (NMY)-2 labeled with green fluorescent protein (NMY-2::GFP) with advancement of an adjacent extension. Fluorescence was normalized to maximal integrated intensity, and extension advancement was normalized to the final position of the tip of the extension in the rosette. (C) Apical contractile flows were required for rosette formation. The panels show a comparison of endoderm covering in (left) an unperturbed embryo and (middle) an embryo in which the apical cortex of Ea and Ep was ablated (lightning) by laser. These panels show three-dimensional (3D) projection still images, and uncovered endoderm is highlighted by a dashed line. (D) Quantification of covering of endoderm in two wild-type embryos and in two embryos in which the endodermal cortices were ablated by laser. (E) A large bleb formed at the ablated cortex. Two-dimensional (2D) confocal slices at higher temporal resolution reveal the site of ablation (dashed circle/lightning), cortex rupture (arrow), and extensions (open arrows).