Fig. 4

(A) Immunoblot analysis and protein quantification showing ALIX/PI3K-C2α expression ratio in zebrafish lens. (B) Immunofluorescence showing (top) ALIX or (bottom) PI3K-C2α expression in 72-hpf zebrafish lens. (C) Immunofluorescence staining showing colocalization between endogenous VPS36 and PI(3,4)P2 at the midbody ring. (D) Snapshots taken from time-lapse imaging of GFP-VPS36 during cytokinesis in HeLa cells stably expressing red fluorescent protein (RFP)–a-tubulin. Arrowheads indicate enrichment of VPS36 at the abscission site. (E) Quantification of VPS36 levels at midbody in control and siRNA-treated cells. n ≥ 200 cells, mean ± SD. (F and G) Immunofluorescence staining and quantification of endogenous (F) VPS36 and (G) PI(3,4)P2 levels at the midbody in fibroblasts derived from patients with homozygous deletion of PI3K-C2α. +/+ genotypes were pulled together. n ≥ 50 cells, mean ± SD. (H) Quantification of CHMP4B levels at midbody in control and siRNA-treated HeLa cells. n ≥ 120 cells, mean ± SD. (I) Quantification of VPS36 levels at midbody in control and siRNA treated cells upon transfection of siRNA-resistant WT, KD, and PI(3)P–producing (C3) forms of PI3K-C2α. n ≥ 30 cells, mean ± SD. If not previously specified, all results are shown as mean or representative picture of at least three independent experiments ± SEM. **P < 0.01; ***P < 0.001.