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Figure 2


Fig. 2. Amnioserosa cell elongation involves rotation of the MT cytoskeleton. (A) Stage 7-8 AS 3D projections. DE-Cad (red). Dlg (green). Purple line and box show a cross-section. Basolateral membranes of two cells are outlined. (B) Three-dimensional projections of Tubulin-GFP as live AS cells elongate. Smooth MT rotation from apical basal axis (0:00, arrows show bundle end-on views) to cell elongation axis (4:24-26:20, arrows). Centrosomes are detected (arrowheads). Gamma adjustments are made to visualize all MTs. (C) Rotation direction quantification. (D) Single rotating cell. Side view. Rotating MT bundles (asterisks). (E,F) Pre-elongation live embryo in single plane surface view (E) or in cross section (F). Tubulin-GFP staining. MT bundles are in cross-section (E, arrow) or seen longitudinally (F, arrow). Centrosomes (F, arrowheads) above elongated nuclei (dark ovals). MTs above centrosomes are bracketed (F). (G,H) During elongation live embryo in single plane surface view (G) or in cross section (H). Tubulin-GFP staining. MT bundles are seen longitudinally (G, arrow) or in cross-section (H, arrow). Centrosomes (G, arrowhead) at one end of elongated nuclei (G, dark ovals). MT extension past centrosomes (G, brackets). (I-K) DE-Cad (red), centrosomes ({gamma}tubulin, green). (I) Pre-elongation, surface view. Centrosomes (arrowhead) above nuclei (inset, cross-section). (J,K) During elongation. Centrosomes at one end of nuclei (arrows, dark circles). (L-N) DE-Cad (red), YFP-ER targeting sequence (green). (L) Pre-elongation, surface view. ER bulk above nuclei (inset, cross-section). (M) During elongation the bulk of the ER is at one cell end (arrows) but evens out later (N). (O,P) Projections. AS cells are at early elongation. Apical Tubulin-GFP (O) and Moesin-ABD-GFP (P). (Q) Pre-elongation, cross-section. DE-Cad, red; acetylated tubulin, green. (R) Elongating, surface view. DE-Cad, red; acetylated tubulin, green; tubulin, green/red. Acetylated microtubules along cell elongation axis (arrows). (S) During elongation, surface view. EB1-GFP staining. Kymogram of box of one cell side. White lines indicate two EB-1 complexes. Scale bars: 5 µm.