Disrupting assembly of the inner membrane complex blocks Plasmodium falciparum sexual stage development
Fig 3
PhIL1 is located at the IMC in gametocytes.
(A) Immunofluorescence microscopy showing anti-PhIL1 (green) at the periphery of a 3D7 stage IV gametocyte, showing fluorescence close to the anti-β-tubulin (red) labeling. (B) Immunofluorescence microscopy of a stage IV gametocyte showing overlap of GAP50-GFP (red) and PhIL1 (green) at the periphery of the cell. (C) Western blot analysis of saponin-treated pellets of 3D7 and GAP50-GFP stage IV gametocytes. Gametocytes were probed with PhIL1 pre-immune serum, anti-PhIL1 antiserum, anti-GAP45, anti-GFP and anti-ERC. (D, E) Immunofluorescence microscopy of stage IV PhIL1-GFP (D) and PhIL1-HA (E) gametocyte transfectants. Parasites were labeled with anti-PhIL1 (green) and anti-GFP or anti-HA (red). Nuclei were labeled with DAPI. Scale bars: 5 μm. (F) Purified, saponin-lysed PhIL1-HA gametocytes were solubilized with different extraction agents for 30 min. Pellet and supernatant fractions were separated and loaded in equivalent amounts on 4–12% acrylamide gels. PhIL1 and GAP45 bands were visualized using mouse anti-HA or rabbit anti-GAP45 primary and HRP-conjugated goat anti-mouse or goat anti-rabbit secondary antibodies, respectively. P = pellet; S = supernatant. Western analysis and immunofluorescence of PhIL1 is presented in S3 Fig.