We found that loss of results in aberrant basal cell extrusion and cell intercalation events. UAS-shRNA and Gap43::CH (membrane). White arrowheads and colored dots track an interface and cells, respectively. (E, F) Schematized interface denoted by white arrowheads in E and F, respectively. (G) Number of cells extruding during tissue folding. (H) Number of T1 transitions during tissue folding. Box plots (in this and all subsequent figures): red line, median; bottom and top, ARHA 25th and 75th percentiles, respectively; black dashed lines, lowest and highest values; red crosses, outliers beyond 1.5 times the interquartile range of the PJ34 box edges. * 0.00001. n.s., not significant. Scale bars, 5 m. See for data point numbers for all experiments in this and all subsequent figures. Abl tyrosine kinase has conserved roles in tissue morphogenesis and disease states (Koleske Abl regulates apical F-actin organization during apical constriction and tissue folding via negative regulation of Enabled (Ena; Fox and Peifer, 2007 ). Ena binds to F-actin barbed ends to promote actin elongation and restrict actin capping (Bear and Gertler, 2009 ; Hansen and Mullins, 2010 ). Abl also promotes AJ dynamics during tissue elongation via -catenin (-cat; gastrulation, ventral cells apically constrict in a coordinated manner; cells constrict their apical surface at similar rates, such that apical surface areas are homogeneous (Figure 1, A and B). Abl is required for this coordinated apical constriction; transcript (Jodoin embryos. Live imaging of or control embryos (Figure 1, BCD, and G, Supplemental Figure S1, E, F, J, and K, and Supplemental Movie S1). Extrusion was not observed in cells adjacent to the ventral region that do not express Twist and Snail (nonventral cells; Figure 1G). This suggests that Abl promotes the maintenance of cells within the epithelium during tissue folding. Loss of results in a disorganized, apical actomyosin meshwork, with some cells lacking apical actomyosin (Fox and Peifer, 2007 ). However, apical actomyosin pulses were observed in extruding cells (Supplemental Figure S1H; 17 of 17 embryos). Nuclei of extruding cells were not fragmented, suggesting that extrusion is not due to an apoptotic signal (Supplemental Figure S1K). Moreover, before the onset of tissue folding, embryos depleted for exhibit reduced cell packing (- 0.00001), suggesting that cell extrusion is not PJ34 due to cell crowding. In addition to extrusion, intercalation events PJ34 known as T1 transitions, in which junctions aligned along the dorsalCventral axis collapse and extend new junctions along the anteriorCposterior axis (Bertet functions to prevent cell extrusion and intercalation specifically in Twist- and Snail-expressing cells during tissue folding. Abl regulates apicalCbasal polarity of ventral cells After tissue folding and tube formation, ventral cells lose apicalCbasal polarity and undergo EMT (Clark depletion PJ34 altered apicalCbasal polarity. During apical constriction, the cell polarity protein Par-3 (depletion resulted in the basolateral accumulation of Par-3 specific to the ventral region (Figure 1A, apically constricting; Figure 2, BCE, red arrows, and Supplemental Figure S1I). This accumulation below that apical surface due to the loss of occurred after the onset of tissue folding (Figure 2C, red arrows). In contrast, Par-3 is restricted apically and not present in the basolateral domain of embryos (Figure 2, A, and CCE, yellow arrows). These data suggest that maintains apicalCbasal polarity in ventral cells during tissue folding. Open in a separate window FIGURE 2: Abl depletion disrupts apicalCbasal polarity in ventral cells. (ACD) Embryos expressing indicated UAS-shRNA and GFP::Bazooka (Par-3). (A, B) Time-lapse images of basolateral domain of ventral cells (21 m below the apical surface). Red arrows denote basolateral Par-3. (A, B) Zoomed-in region indicated by the white-dashed boxes in A and B, respectively. Red arrows denote dynamic basolateral Par-3. (C) Kymographs of embryos expressing indicated UAS-shRNA and Par-3. Kymographs of basolateral line along the anteroposterior axis. Red arrows denote basolateral Par-3, and blue arrowhead indicates the beginning of tissue folding. (D) Cross-sections. Yellow arrows denote apical Par-3, and red arrows denote basolateral Par-3. (E) Ratio of ventral vs. nonventral basolateral Par-3 intensity. (F) Fixed images of the apical domain in embryos expressing the indicated UAS-shRNA stained for E-cad. Apical E-cad, AJs (green). Subapical E-cad, cell outline (gray). White arrowheads denote pools of medioapical E-cad. (G) Fixed images of the basolateral domain (21 m below the apical surface) in embryos expressing the indicated UAS-shRNA and stained for E-cad and phalloidin (F-actin). White arrows denote junctions lacking E-cad, and white arrowheads denote junctions overaccumulated with E-cad. (H) Basolateral E-cad intensity, coefficient of variance. (I) Cross-sections of fixed embryos expressing.