Arrangement of mitochondrial nucleoids during life cycle of Saccharomyces cerevisiae.

The fluorescent dye, 4',6-diamidino-2-phenylindole (DAPI) intensely stains cytoplasmic DNA in plant (1) and yeast cells (2, 3).In Saccharomyces yeast cells, mitochondrial DNA (mtDNA) was observed as a bright nucleoid "chondriolite" (2) and with a "string-of-beads" appearance (3) during growth.On the other hand, the role and behavior of mtDNA might be attractive in relation to nuclear division during meiosis of Saccharomyces cerevisiae, since active functioning of mitochondria is essential for sporulation (4-6).We report here that mtDNAs aggregate and form structures with a "string-of-beads" appearance and then separate again into individual "chondriolites" during the meiosis-sporulation and cell division cycles.S. cerevisiae strain G2-2 was used (7).Presporulation culture was performed in modified Burkholder's medium, which was the same as that used by SANDO and MIYAKE (7), except for the omission of amino acids and purine and pyrimidine bases and supplementation with tomato-juice (3.6 %), under reciprocal shaking at 30°.Cells at the stationary phase were harvested, and small age-0 cells with no bud scar (Sm) or large cells of age one or above with bud scar (s) (La) were separated by repeated differential and density gradient centrifugations, using sorbitol or dextran (lot No. YH-5670, supplied by courtesy of Meito Sangyo Co., Ltd.).For sporulation culture, the La cells were inoculated into sporulation medium consisting of 60 mM potassium acetate and 50 mM potassium phosphate (pH 6.89), and incubated at 30° under reciprocal shaking.Ninety-one percent of La cells sporulated within 48-hr incubation.Synchronous culture was obtained by inoculating the La cells into the presporulation medium and incubation at 30° under reciprocal shaking.Aliquots from the cultures were sampled at regular time intervals and the cells were 511 5.12 SANDO, MIYAKAWA, NISHIBAYASHI, and KUROIWA VOL.27