Cubilin/AMN-mediated protein uptake is essential for maintaining subcellular structures of Drosophila nephrocytes. (A–C) The wild-type (WT) Drosophila nephrocytes show distinct layers of (A and B) lacuna network and endocytic vacuoles and lysosomes. (C) Nephrocyte diaphragms and lacunae densely populate the nephrocyte surface. (D–F) Pericardial nephrocytes with dCubilin RNAi knockdown have (D and E) fewer endocytic vesicles and vacuoles and almost no electron-dense lysosomes. (F) The number of nephrocyte diaphragms is reduced, and the lacunae become much shorter. (G–I) dAMN RNAi knockdown also reduced the number of (G and H) endocytic vacuoles and electron-dense lysosomes, with (I) a reduced number of nephrocyte diaphragms and shorter lacunae. There is no dramatic change of basement membrane. Scale bars, 4 μm in A, D, and G; 1 μm in B, E, and H; 200 nm in C, F, and I. (J) Quantification of the ultrastructure changes in nephrocytes with dCubilin or dAMN RNAi. ND, nephrocyte diaphragm; EV, endocytic vacuole; Lyso, lysosomes.