Fig. 4

Clathrin accessory proteins for assembly (Lap) and disassembly (Aux; Hsc70-4) are required for endocytosis of slit diaphragm protein Pyd. Immunofluorescence labeling for Pyd (red), with DAPI (blue) nuclear stain of adult fly nephrocytes, visualized by confocal microscopy. Scale bar: 1 μm. a–a″ In wild type (Control) nephrocytes, Pyd was localized to the cell membrane (a) and exhibited a uniform and smoothly distributed fingerprint-like pattern on the cell surface (a′–a″). b–b″ Silencing of lap resulted in intracellular aggregation of Pyd proteins with partial disruption of Pyd cell surface localization. Note the fingerprint-like pattern remained largely intact, however gaps between adjacent Pyd-positive ridges appeared increased. c–d Silencing of aux (c–c″) and Hsc70-4 (d–d″) resulted in severe disruption of Pyd cell surface localization. Revealing intracellular localization of aggregate-like structures, and cell-surface distribution marked by high-density aggregation areas