Fig. 3.

Loss of the E-hook attenuated IFT of GFP–β-tubulin. (A) Schematic presentation of tubulin binding by the IFT74N–IFT81N module (as proposed by Bhogaraju et al., 2013). (B) The frequency of anterograde IFT events observed for full-length (WT) and truncated (ΔE-hook) GFP-tagged β-tubulins in cilia of the ift81-1 IFT81 and the ift81-1 IFT81-CH5E strains. The number of regenerating cilia analyzed for each strain (n) is indicated. Data are mean+s.d. The corresponding western blot comparing the amounts of wild-type and ΔE-hook GFP–β-tubulin in whole-cell samples of the strains is shown below the graph. Tagged tubulin was detected with anti-GFP, anti-NAB1 staining was used to verify equal loading. (C,D) Anterograde transport of full-length and E-hook-deficient GFP–β-tubulin (C) and GFP–α-tubulin (D) in the ift74-2 IFT74Δ130 strain and wild-type (WT) controls. Data are mean+s.d., and n for each strain is indicated. Western blots of whole-cell samples documenting similar expression of the transgenes are below. (E) Frequency of anterograde IFT for full-length and E-hook-deficient GFP–α-tubulin in the ift81-1 IFT81 and the ift81-1 IFT81-CH5E strains. Data are mean+s.d., and n for each strain is indicated. The corresponding western blots of the whole-cell samples of these strains are shown below the graph. (F) Anterograde transport of αE-hook–GFP–β-tubulin in the ift81-1 IFT81 and the ift81-1 IFT81-CH5E strains. Data are mean+s.d., and n for each strain is indicated. The corresponding western blots of the whole cell samples are shown. Diagrams in B–F depict the mutations in tubulin, IFT74 and IFT81 in each strain.