Abnormal BB localization in ta3−/− PR underlies the OS development defect. (A,B) 4 dpf cryosections stained with BODIPY (red) marking membranes of the OS and mitochondrial cluster and with anti-Centrin antibody (green) to mark the basal bodies, show aberrant localization of BB below the mitochondria in ta3−/− PRs. In wildtype (wt) PRs, the centrin-marked BB (arrow in A’) is located just basal to the OS (straight bracket) and apical to the mitochondrial cluster (curved bracket), while this localization is lost in a substantial number of ta3 mutant PRs with BBs located below the mitochondrial cluster (arrow in B’). (C,D) Immunofluorescence on 4 dpf cryosections with anti-VDAC1 antibody (red) to mark the mitochondria and anti-Centrin antibody to mark the BB showing aberrant positioning of BBs in ta3 mutants basal to the mitochondria (arrows in D) compared to wt (C). (E) Quantification of BB position with respect to the mitochondrial cluster based on immunofluorescence with anti-Centrin antibody. Each data point represents the proportion of PRs with BBs below the lower 1/3 of the mitochondrial cluster on one confocal section of a whole retina from one single larva. The proportion of 4 dpf PRs with aberrant BB positioning below the mitochondrial cluster is significantly increased in ta3 mutants (****p < 0.0001, Student’s t-test, n = 10 wildtype and 9 mutant larvae). Bars are standard deviation. (F–F”) Representative TEM images showing presence of a BB in cross-section (arrow) within a mitochondrial cluster even at the later stage of 5 dpf, when all BBs should have docked to the apical membrane. (F’) Is the boxed area in (F) and (F”) is the boxed area in (F’). BB basal body, IS inner segment, m mitochondria, N nuclei, ONL outer nuclear layer, OS outer segment, PR photoreceptor. Scale bars: 10 µm in (A,B), 3 µm in (A’–B’), 4 µm in (C,D), 3 µm in (F), 1 µm in (F’) and 0.5 µm in (F”).
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