Fig. 1: Quantitative analysis of Tribolium serosa expansion.

a Schematic depiction of the geometric constraints experienced by a tissue expanding over a spherical yolk cell. The leading edge undergoes an area increase followed by an area decrease after it crosses the equator. b Illustrations of the stages of Tribolium embryogenesis from cellular blastoderm to serosa window closure. c 3D renderings of a Tribolium embryo expressing the fluorescent H2A-eGFP nuclear marker reconstructed from a multi-view time-lapse SPIM recording. The embryo is shown from the lateral and ventral views at the six reference stages corresponding to the schematics in b. All imaged embryos in this and other panels are shown with anterior to the left, and all time stamps are in hh:mm. Scale bar is 50 µm. N = 1 (2 datasets available). d 2D cartographic projections at reference stages of a 4D SPIM recording of a Tribolium embryo expressing EFA-nGFP. The extent of the serosal tissue is highlighted in turquoise. Scale bar is approximately 100 µm (see “Methods”). N = 1. e The area of the serosal tissue calculated from cartographic projections of 4D SPIM recordings. The data are normalized to the total serosa area at Stage 5 in each case. For every stage, the total serosa area is calculated for all time points between two consecutive stages in three different embryos and plotted as a distribution. Plots in this and all other panels indicate the median with a thick line, the mean with a black dot, and the standard deviation (s.d.) with the thin error bars. N = 3. f Comparison of the distributions of apical areas of cells sampled from confocal recordings of Tribolium embryos expressing the cortical LifeAct-eGFP actin marker at reference stages labeled according to b. The number of cells (n) and the number of embryos (N) sampled at different stages were in the dorsal region Stage 0 n = 58 and N = 6, Stage 1 n = 116 and N = 11, Stage 2 n = 66 and N = 9, Stage 3 n = 39 and N = 6, Stage 4 n = 76 and N = 10, and in the ventral region Stage 3 n = 52 and N = 7. The normal distribution of the data was tested using Shapiro–Wilk test. Distributions were compared using the non-parametric two-sided Wilcoxon Rank-Sum test (same in all figures unless stated otherwise). p Values between 0.05 and 0.01 are labeled with single asterisk (*), 0.009–0.001 are labeled with double asterisks (**), <0.001 with triple asterisks (***), and ns signifies a non-significant p value (same in all figures). g Cartographic projections at reference stages of a transgenic embryo labeled with LifeAct-eGFP and reconstructed from a multi-view SPIM recording. All serosal cell in each projection were segmented automatically, curated manually, and color coded according to their apical cell area. Red boxes indicate the approximate regions from which cells sampled in confocal datasets were quantified in f. N = 1.