Isolation of a taxol-resistant Chinese hamster ovary cell mutant that has an alteration in α-tubulin

Abstract

Taxol is a plant alkaloid that has antimitotic activity and appears to stabilize microtubules. Taxol-resistant cells were selected from a population of UV-mutagen-treated Chinese hamster ovary cells by a single-step procedure. These mutants have normal morphologies and growth rates but are 2- to 3-fold more resistant to the toxic effects of the drug than the wild-type parent. One out of 20 mutants screened by two-dimensional electrophoresis for chemical alterations in tubulin had an 'extra' spot with a more acidic isoelectric point than α-tubulin. This extra spot was shown to be an electrophoretic variant α-tubulin by its copurification with tubulin in crude microtubule-containing preparations and by one-dimensional peptide mapping. The α-tubulin mutant was found to be temperature sensitive for growth, and this property was used as the basis for the selection of revertants. Seventeen temperature-resistant revertants of the α-tubulin mutant were selected for their ability to grow at 40°C and three of these revertants were found to have simultaneously lost their taxol resistance and the electrophoretic variant α-tubulin. These results provide evidence that an alteration in α-tubulin can confer taxol resistance on a mammalian cell line and suggest that α-tubulin is essential for cell viability.