Exocytosis is inhibited before metaphase-anaphase transition. (A) The cdc20-1 mutant exhibits Bgl2 secretion defects at the restrictive temperature. Cells were grown at 25°C or shifted to 37°C for 1.5 h. Internal (In) and external (Ex) pools of Bgl2 in wild-type, cdc28-4 (G1 phase), cdc7-1 (S phase), cdc20-1 (metaphase), and cdc15-2 (anaphase) cells were examined by Western blot analysis. Alcohol dehydrogenase (ADH) levels were probed as a protein loading control. (B) The internal accumulation of Bgl2 was quantified for the conditional cell cycle mutants. The stages of cell cycle arrest for these cells are indicated: Meta, metaphase; Ana, anaphase. AU, arbitrary units. Error bars represent standard deviation (n = 3). *, P < 0.01. (C) Selective block of glycoprotein secretion in the cdc20-1 mutant at the restrictive temperature. The asterisks indicate the bands with reduced levels of secretion in the cdc20-1 mutant. (D) Quantification of the levels of protein indicated by the arrow in C. Error bars represent standard deviation (n = 3). *, P < 0.01. (E) Thin-section EM shows that cdc20-1 cells accumulate post-Golgi secretory vesicles (typically 80-100 nm in diameter) at the restrictive temperature. “SV” and the arrow indicate one of the vesicles. Bars, 1 µm. (F) Quantification of vesicle numbers per EM section detected in the wild-type and cdc20-1 cells. Error bars represent standard error of the mean (n = 35). *, P < 0.01.