Tubulin isotypes in budding yeast are not functionally equivalent. (a) Genetic organization and viability of yeast cells harboring deletions of endogenous α-tubulin isotypes. (b and c) Representative Western blot (b) and quantification from three independent experiments (c) of α-tubulin levels in WT and tub3Δ cells. Actin is loading control. Mean ± SEM; ***, P ≤ 0.001 by unpaired, two-tailed Student’s t test. Diamonds show individual values from each experiment. (d) Viability of cells with a single α-isotype gene under regulation of the other isotype locus. (e) Tetrad analyses from diploids with the indicated deletions and substitutions of TUB1 and TUB3 genes. ⁺, viable spores; ⁰, inviable spores. (f) Representative tetrad dissections of strains from panel e. Sister spores are arranged vertically. Green boxes mark representative spores of the indicated genotype to show growth is comparable to sister spores. No haploid spores of tub1Δ::TUB3 tub3Δ::URA3 genotype were viable (far right). Red boxes mark representative inviable spores that, based on genotype of their surviving sisters, harbor the tub1Δ::TUB3 tub3Δ::URA3 genotype. As reported, diploids heterozygous for tub1Δ or tub3Δ also display overall decreased spore viability relative to WT (Schatz et al., 1986b).