Fig. 6

Rapamycin treatment promotes autophagy in gja8b mutant zebrafish lenses. (A-C) Electron micrographs of lens fiber cells (LFC) and lens epithelial cells (LEC) in 72 hpf WT, gja8b mutants, and gja8b mutants treated with 25 μM rapamycin. Yellow arrowheads show AV. (B) The number of AVs per area (40 μm²), and (C) is the average area of AVs in the lens epithelial and fiber cells in WT, gja8b mutants and gja8b mutants treated with 25 μM rapamycin (n ≥ 60 cells from 6 lenses). (D) Shows the concentration of endogenous Map1lc3b-I and Map1lc3b-II in the zebrafish eyes of WT, gja8b mutants, WT treated with 25 μM rapamycin and gja8b mutants treated with 25 μM rapamycin. Short exp and long exp: short exposure and long exposure. (E) shows the quantitative analysis of relative intensity of Map1lc3b-II normalized to the Tubg1 loading control (n = 3 independent experiments). (F) Shows the concentration of endogenous GJA8 and LC3 in NC or rapamycin-treated HLE cells. (G) Shows the quantitative analysis of relative intensity of GJA8 and LC3-II normalized to the TUBG loading control (n = 3 independent experiments). (H) Shows representative images for endogenous ATG16L1 and GJA8 in NC and rapamycin treated HLE cells. (I) Quantitative analysis of the average number of colocalized GJA8 and ATG16L1 puncta per cell. (n = 3 wells, 3 independent experiments, > 50 cells per experiment). Scale bar: 100 nm (A) and 10 μm (H). Mean ± SEM, N.S., p > 0.05, *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001