Ventral furrow formation was inhibited upon photo-stabilization of myosin-II levels at the cell base. Embryos co-expressing the optogenetic module and the myosin-II probe Sqh::mCh were mounted with the ventral tissue facing the objective. A region of photo-activation (two-photon λ=950 nm) was defined at the cell base in the anterior half of the embryo, and whole-cell myosin-II signal was recorded (confocal setup; λ=561 nm) in an alternating fashion. Scale bars, 20 µm. (A) Top view showing apical myosin-II 10 min after the initial activation. While cells in the non-activated region have internalized and disappeared from the apical focal planes (right), cells in the activated region failed to internalize despite showing myosin-II accumulation at the apical surface (left). Red and Blue dotted lines indicate the location of the cross-section panels displayed in (C) and (D). (B) Quantification of apical myosin-II levels demonstrate that the defects in tissue invagination were not due to impaired apical myosin-II accumulation as the average apical myosin-II level per cell did not significantly differ between the non-activated (NR) and activated region (AR). The intensity values were normalized using the median value of the combined cell population in the NR and AR. (N = 3 embryos; nNR: 94, nAR: 78). Significances (by two-sample t-Test): n.s. indicates no statistically significant differences. In each box plot, the central mark, the bottom, and the top edge of each box indicates the median, the 25th and 75th percentile, respectively. Whiskers extent to the most extreme data point and the '+' symbol indicates an outlier. (C 