Direct involvement of the isotype-specific C-terminus of tubulin in ciliary beating

In previous studies in Drosophila, Nielsen et al. hypothesized that the tubulin C-terminal axonemal motif 'EGEFXXX', where X is an acidic amino acid, is required for ciliary function and assembly (Nielsen et al., 2001, Curr. Biol. 11, 529-533). This motif is present in some but not all mammalian tubulin isotypes. We therefore investigated whether this motif is important in ciliary function in mammals. In a preparation of isolated, ATP-reactivated bovine tracheal cilia, we found that monoclonal antibodies directed against the C-terminus of _I, _IV and _V tubulin blocked ciliary beating in a concentration dependent manner. Antibodies against other epitopes of tubulin were ineffective, as were antibodies against tubulin. Peptides consisting of the axonemal motif and motif-like sequences of these isotypes blocked ciliary beating. These results suggest that the axonemal motif sequences of _I, _IV and _V tubulin are essential for ciliary function. Peptides consisting of corresponding C-terminal sequences in tubulin isotypes were also ineffective in blocking ciliary beating, which suggests that the C-terminus of tubulin is not directly involved in cilia function in mammals.