Table II.

Suppression of kar3-Δ Mitotic Delay by Treatments that Destabilize Microtubules

Genotype		Cells in mitosis (anti-spb staining)		Cells arrested in mitosis (DAPI staining)
Wild-type		24% ± 5.4 SD		4.7% ± 1.6 SD
kar3-Δ		46% ± 7.1 SD		34% ± 5.2 SD
kar3-Δ tub3-Δ		31% ± 9.6 SD		7% ± 3 SD
Wild-type + nocodazole		25% ± 6.9SD		11.5% ± 2.9 SD
kar3-Δ + nocodazole		29% ± 6.6 SD		10% ± 4.4 SD
tub3-Δ		22% ± 4.2 SD		2.2% ± 0.8 SD

As observed previously (Meluh and Rose, 1990), loss of KAR3 produced an increase in the percentage of cells in the preanaphase mitotic phase of the cell cycle, suggesting a delay in initiation or completion of nuclear division. Unsynchronized, log phase cells grown at 30°C were fixed and stained with anti-spindle pole antibodies and DAPI and examined by epifluorescence microscopy. The percentage of cells with separated spindle poles as determined by anti-spb staining and with large buds and a single nucleus as determined by DAPI staining is shown. Cells with separated spindle pole bodies were considered to be in mitosis. Large budded mononuclear cells were considered to be delayed or arrested in mitosis. The increase in mitotically arrested kar3 mutants was mostly or completely corrected if the TUB3 gene was deleted or if 3.3 μg/ml nocodazole was added to the culture. Addition of nocodazole to the wildtype population resulted in a slight increase in mitotically arrested cells, mostly likely indicating a checkpoint arrest response to this microtubule inhibitor. Large buds were defined as greater than ∼50% the size of the mother. 200–400 cells/sample were counted on each of three separate trials; the average is given.