Fig. 2: Cells undergoing tissue-folding events during Drosophila gastrulation show high Brillouin shifts.
a, Surface view of a fixed Drosophila embryo undergoing VFF (arrow). The embryo expresses the membrane marker Gap43-mCherry to visualize cell outlines. The green arrowhead indicates the direction of Brillouin illumination. The red dashed line indicates the approximate location of the physical cross-sections in b and c. b, Stages of VFF progression shown in cross-sections of fixed embryos, stained with a Snail antibody (green) to label the nuclei of cells engaged in VFF and a disks large antibody (magenta) to label cell outlines. c, Top: SPIM images of an embryo expressing Gap43-mCherry to mark cell membranes; bottom: median-projected Brillouin shift maps from the same embryo at the same timepoints and positions (representative embryo of n = 3 in total). The arrow indicates the formation of the ventral furrow. The dashed box encloses the potential mesoderm region in the Brillouin shift maps used for quantification in d. d, Quantification of Brillouin shift averaged over 18 cells engaged in apical constriction and invagination during VFF progression, as inferred from SPIM slices (dashed box in c). Changes in Brillouin shift are shown relative to the Brillouin shift at timepoint 0 min, which corresponds to the initiation of apical constriction. Data points are average Brillouin shift in the contractile domain, error bars denote s.d. over different slices of each analyzed embryo. Statistical significance was evaluated using a paired one-way ANOVA test (F = 44.29, P = 0.066), followed by a post-hoc multiple comparison test (false discovery rate corrected, α = 0.05, q-value = 0.1); *P = 0.039, **P = 0.0054 (n = 3 embryos from independent experiments). e, PMI in embryos at a similar stage as in a. Top: mid-sagittal section; inset: higher magnification of the posterior midgut region showing apical constriction and invagination of the posterior midgut primordium; bottom: surface dorsal view of a Drosophila embryo showing the formation of the circular contractile domain of the posterior midgut that encloses the pole cells (arrow). f, Representative SPIM images (top) and median-projected Brillouin shift maps (bottom) at five timepoints during invagination (out of n = 3 in total). The arrow indicates the formation of the circular contractile domain, comparable to the PMI progression in e (inset). The arrowhead indicates the displacement of the posterior end during PMI. The initiation of apical constriction was set as the 0 min timepoint. Images in Gap43-mCherry and Brillouin panels are median projections of three slices of the re-sliced region of interest (see Methods). A, anterior; D, dorsal; P, posterior; V, ventral. Scale bars, 50 µm.
