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Fig. 4. Two indispensable domains for primary-cilium targeting exist in the C-terminus of Inv. (A) Primary-cilium localization of GFP-tagged truncated forms of C-terminal Inv was observed by GFP fluorescence and DIC in living cells. Truncated Inv constructs were introduced into the inv cell line. Numbers indicate the position of amino acids. The wedged marking indicates the deletion of the IQ2 domain. Arrowheads in the images indicate the GFP signal and primary cilia. Primary cilia in living cells are seen as a dot. Results of the localization of GFP-tagged truncated Inv are summarized on the right. –, Inv absent; +, Inv present; +/–, part of the transfected cells shows ciliary localization of truncated Inv. Percentages of ciliated cells with positive ciliary GFP signal in transfected cells are also indicated. At least two independent regions, amino acids 915-935 (blue box: the IQ2 domain) and amino acids 1030-1062 (red box), are necessary for cilium targeting. (B) Localization of Inv (842-1062) in the primary cilia was further confirmed by immunocytochemistry. The primary cilium was detected by anti-acetylated -tubulin antibody (ac-tubulin; red). Inv (842-1062) signal was predominantly detected at the base of the primary cilium. Scale bars: 10 µm.

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