Figure 2.
tba-6 Is Required for CEM Cilia Curvature, Microtubule Architecture, and Tubulin Composition(A-B') Serial TEM reconstructions of male cephalic sensilla of WT and
tba-6 males. Pie charts (bottom) indicate the penetrance of the cilia curvature phenotype asassayed by TBB-4::tagRFP (WT N = 83,
tba-6 N = 102 males).(A) In WT, CEM (yellow) and CEP cilia (gray) share a lumen formed by the sheath (inner border in pink) and socket (teal) (refer to Figure 1A). The distal tip of the CEMcilium protrudes to the environment through a cuticular pore (brown) (also Figure S1C). The CEP cilium curves in and embeds in the cuticle. EVs (green) are locatedin the proximal region of the cephalic lumen.(B-B') A total of 59% of
tba-6 cilia curve-out but do not protrude through the cuticular pore. EVs were observed inside the cuticular pore (for TEM cross-sections oftba-6, see Figure 4C). (B') A total of 41% of
tba-6 cilia curve in, follow the path of the CEP cilium, and embed in the cuticle. EVs were found in the lumen surrounding CEM and CEP cilia.(C) In
tba-6, microtubule doublets terminate and do not form A- and B-tubule singlets. Right: serial TEM images of
tba-6 cilia with A-tubules continuing as singlets and B-tubules terminating (black arrow). Left: cross-sections of WT cilia from a serial tomogram showing a representative A- and B-tubule of a microtubuledoublet splitting to form two singlets (white arrow) (refer to Figures 1F, S1A, and S1B). Scale bar, 50 nm.(D) Cartoon depicting microtubule architectures and cilia shapes. (E) Cross-sections of sub-distal CEM cilia in WT and
tba-6 animals. Scale bar, 50 nm. Right: microtubule singlet quantification: WT = 20 +- 2,
tba-6 curved-out =14 +- 2 tubules, mean +- SD, p value = 0.0269 by Mann-Whitney, N = 3 animals, and n = 12 cilia for each genotype.(F-G') b-tubulin TBB-4::GFP relative ciliary levels and ciliary localization. In WT, TBB-4::GFP localizes to CEM cilia in a stereotypical pattern; lines indicate two offour CEM cilia, quantified in (G), n = 25 cilia. Ciliary TBB-4::GFP levels were normalized to levels in the CEM dendrite, mean fold intensity +- SD. This pattern isaltered in
tba-6 mutants, quantified in (G0), n = 20 cilia. In
tba-6, TBB-4::GFP ciliary levels are significantly reduced and TBB-4::GFP accumulates at ciliary bases.(F0) N = 86 and 102 animals for WT and
tba-6 respectively, p value < 0.0001 by Mann-Whitney U test, Scale bar, 10 um.See also Figure S2 and Table S1.