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Fig. 2. Immunohistochemical analysis of wild type and IFT mutants. Retinal cryosections of 4 dpf larvae were stained with 1D1, an antibody that labels rhodopsin, with BOPS, an antibody against blue cone opsin, and with ZPR1, an antibody that recognizes an unknown epitope of red-green double cones. In addition, anti-acetylated tubulin (AT) was used to label microtubules and visualize cilia. In all panels, immunolabel is shown in green and nuclei are counterstained with DAPI (blue). (A) Wild-type photoreceptors have rhodopsin localized almost exclusively to the outer segment (arrowhead). (B) IFT57 mutants display significant rhodopsin localization to the outer segment (arrowhead), but punctate areas of rhodopsin mislocalization (inset, red arrowhead) are observed, as well as mislocalization of rhodopsin through the plasma membrane (red arrow). (C) IFT88 mutant photoreceptors have rhodopsin completely mislocalized throughout the plasma membrane (red arrow) and both IFT mutants exhibit cell death in the photoreceptor layer, as indicated by condensed and bright DAPI-labeled nuclei of pyknotic cells (white arrow). (D-F) Blue opsin also localizes to the outer segments of blue cones. IFT57 and IFT88 mutants exhibit a similar pattern of mislocalization with blue cone opsin as seen with rhodopsin. (G-I) ZPR1 labeling indicates red-green double-cone morphology at 4 dpf in wild type. IFT57 and IFT88 mutants have shorter cones when measured in the apical-basal axis, and adopt an abnormal morphology. (J-L) Anti-acetylated tubulin stains microtubules in the connecting cilia (arrowheads) that project apically from the cell body in both wild type and IFT57 mutants, but not in IFT88 mutants. Scale bar: 10 µm.
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