α-Tubulin is a substrate of the ubiquitin conjugation system in flagella. (A–D) Wild-type (WT), fla10, fla8, and fla3 cells were shifted to 32°C for 1 and 1.5 h (as indicated on top of the blots), and immunoblots of flagella from these cells were probed with antibodies to α-tubulin (A), acetylated α-tubulin (B), β-tubulin (C), and IC2 (D). (E) Anti–α-tubulin antibody was used to immunoprecipitate tubulin from an extract of flagella isolated from fla10 after shifting to 32°C for 1 h. Immunoblots of the precipitate were probed for α-tubulin (α-tub) and ubiquitin (Ub). The 50-kD bands include the heavy chain of the precipitating IgG. Molecular masses are given in kilodaltons.