He colR4 and colR15 β-tubulin mutations in Chlamydomonas reinhardtii confer altered sensitivities to microtubule inhibitors and herbicides by enhancing microtubule stability

Abstract

The colR4 and colR15 β2-tubulin missense mutations for lysine-350 in Chlamydomonas reinhardtii (Lee and Huang, 1990) were originally isolated by selection for resistance to the growth inhibitory effects of colchicine. The colR4 and colR15 mutants have been found to be cross resistant to vinblastine and several classes of antimitotic herbicides, including the dinitroanilines (oryzalin, trifluralin, profluralin, and ethafluralin); the phosphoric amide amiprophos methyl; and the dimethyl propynl benzamide pronamide. Like colchicine and vinblastine, the antimitotic effects of these plant-specific herbicides have been associated with the depolymerization of microtubules. In contrast to their resistance to microtubule-depolymerizing drugs, the mutants have an increased sensitivity to taxol, a drug which enhances the polymerization and stability of microtubules. This pattern of altered sensitivity to different microtubule inhibitors was found to cosegregate and corevert with the β-tubulin mutations providing the first genetic evidence that the in vivo herbicidal effects of the dinitroanilines, amiprophos methyl, and pronamide are related to microtubule function. Although wildtype like in their growth characteristics, the colR4 and colR15 mutants were found to have an altered pattern of microtubules containing acetylated α-tubulin, a posttranslational modification that has been associated with stable subsets of microtubules found in a variety of cells. Microtubules in the interphase cytoplasm and those of the intranuclear spindle of mitotic cells, which in wild-type Chlamydomonas cells do not contain acetylated α-tubulin, were found to be acetylated in the mutants. These data taken together suggest that the colR4 and colR15 missense mutations increase the stability of the microtubules into which the mutant β-tubulins are incorporated and that the altered drug sensitivities of the mutants are a consequence of this enhanced microtubule stability.