Fig. 4

Ca²⁺ influx induced by cyclic activation of opto-cCRAC enhances feather elongation. a In Hela cells opt-cCRAC induced Ca²⁺ influx after blue light exposure (n = 20/20). jRCaMP1b was expressed to detect Ca²⁺ level changes. Arrowheads in the line plot highlight the time point of images on the left. F/F₀, changes in fluorescence. Mean ± s.d. Scale bar, 10 µm. b Light-induced Ca²⁺ influx occurred in H&H 35 mesenchymal cells infected by virus encoding opto-cCRAC (cyan rectangles, n = 25/43) but not in the uninfected cells (magenta rectangles, n = 0/33). Mean ± s.d. Scale bar, 10 µm. (′′ denotes seconds). F_E: fluorescence at the end of recording (70′′); **P < 0.01 (Chi-square test). c After 48 h cyclic blue light illumination, skin regions (n = 8/10) highly expressing opto-cCRAC developed more elongated feathers than the poorly infected buds (n = 60 for each group, **P < 0.01, Wilcoxon rank test). No discernable feather morphology changes (n = 48, NS, not significant, Wilcoxon rank test) were observed in skins infected by RCAS-GCaMP6s-T2A-mCherry. Regions close to the dorsal mid-line were magnified (rectangles) to compare feather aspect ratio (ellipses). Scale bar, 500 µm. Customized boxplot: Mean (red) ± s.d. (pink), 95% confidence interval (violet). Dots denote individual data points