Past1 and Syx16 control PI4KII tubulation and PI4P accumulation on SG membranes. (A–E) Spinning-disc confocal images of live L3 salivary gland cells. GFP-PI4KII (grayscale) localization in control (A), Past1^110-1 (B), and Syx16 RNAi (C) salivary gland cells. GFP-PI4KII tubules are marked by yellow arrowheads. (D and E) Still images of GFP-PI4KII tubule dynamics tracked by yellow arrowheads in control (D) and Past1^110-1 mutant (E). See Video 1 for the original videos. (F) Average velocity of the tips of GFP-PI4KII tubules in control and Past1^110-1 mutant. n = total of five tubules from two independent salivary glands per genotype; **, P < 0.01. Error bars indicate standard deviation. (G and H) Leica point-scanning confocal images of live late L3 salivary gland cells processed with Leica Lightning algorithms. Cells expressing Sgs3-GFP (green) and mCh-2xP4M (red) in control (G) or Past1^110-1 mutant (H) SGs coated by mCh-2xP4M are marked by magenta arrowheads. mCh2-xP4M endosome lacking Sgs3-GFP is marked by yellow arrowhead.