Supplementary Figure 6: Wild-type and mutant Skywalker-GFP traffic to and are present at synaptic terminals
From: Skywalker-TBC1D24 has a lipid-binding pocket mutated in epilepsy and required for synaptic function
(a) Confocal images of the ventral nerve cord in third instar Drosophila larvae expressing wild type GFP-Sky (SkyWT) or mutant GFP-Sky (SkyR79C, SkyR281C or Sky3Glu). Scale bar for all top panels: 20 μm.
(b) Confocal images of larval neuromuscular junction endplates of animals with genotypes as in (a). (n = 6 animals), scale bar for all panels in (b): 20 μm. GFP intensities between the GFP-Sky mutants is not significantly different.
(c) Quantification of third-instar NMJ GFP intensity at the membrane in single confocal sections. Mean Fluorescence Intensity between the genotypes GFP-SkyWT, GFP-SkyR79C and GFP-Sky3Glu is similar. Error bars: mean ± s.e.m. (n = 6) P = 0.446, ns: not significant by ANOVA, Dunnett’s.
(d) Quantification of Western blot band intensity of GFP-Sky and GFP-Syt in protein isolations from adult fly heads. Intensities are normalized to Syntaxin control levels. GFP-Sky WT and mutant levels are similar. Error bars: mean ± s.e.m. (n = 3) P = 0.681, ns: not significant by ANOVA, Dunnett’s. The levels of Synaptotagmin-GFP (Syt) are shown as a control.
