Vacuole Segregation in theSaccharomyces cerevisiae vac2-1 Mutant: Structural and Biochemical Quantification of the Segregation Defect and Formation of New Vacuoles

Abstract

The conditional vacuolar segregation mutant vac2‐1 [Shaw and Wickner (1991) EMBO J. 10,1741–1748] shifted to non‐permissive temperature (37°C), forms large‐budded cells without a vacuole in the bud, and daughter cells without an apparent vacuole. Some cells still contain normal segregation structures. Structural and biochemical quantification of the segregation defect showed that (i) about 10% of the full‐grown buds did not contain a vacuole, (ii) about 15% of the small cells washed out of a population growing in an elutriation chamber at 37°C, did not contain a visible vacuole, and (iii) 15% of the cells per generation lost carboxypeptidase Y activity after proteinase A depletion. Thus, 10–15% of the daughter cells did not inherit vacuolar structures or vacuolar proteolytic activity from the mother cell. To investigate the fate of vacuole‐less daughters, these cells were isolated by optical trapping. The isolated cells formed colonies on agar plates that consisted of cells with normal vacuoles, both at 23 and 37°C. Thus, the vacuole‐less cells that failed to inherit proteolytic activities from the mother cell apparently give rise to progeny containing structurally normal vacuoles. Time‐lapse experiments showed that vacuole‐less daughter cells formed vacuolar vesicles that fused into a new vacuole within 30 min. Although new buds only emerged after a vacuole had formed in the mother cell, the temporary lack of a vacuole had little effect on growth rate. The results suggest that an alternative pathway for vacuole formation exists, and that yeast cells may require a vacuole of some minimal size to initiate a new round of budding. © 1997 by John Wiley & Sons, Ltd.

Pex3 peroxisome biogenesis proteins function in peroxisome inheritance as class V myosin receptors

“…overexpressing Pex3Bp activates de novo peroxisome biogenesis in the empty buds and mother cells, respectively (Videos 2 and 4). This is similar to what is observed in mutants of vacuole inheri tance in which buds without vacuoles are rapidly able to form new vacuolar structures de novo, thereby allowing the bud to develop and go on to produce daughter cells of its own (Weisman et al, 1987;Raymond et al, 1990;Gomes De Mesquita et al, 1997).…”

A Role for Actin, Cdc1p, and Myo2p in the Inheritance of Late Golgi Elements inSaccharomyces cerevisiae

“…A portion of the mitochondrial and vacuolar material present in the mother cell is transferred into the bud. However, when vacuolar transfer is inhibited, the daugh-ter cells still acquire a vacuole, probably via the fusion of Golgi-derived vesicles with endocytic vesicles (Hill et al, 1996;Gomes De Mesquita et al, 1997). This type of de novo formation of compartments may be a general feature of the exocytic and endocytic pathways (Glick and Malhotra, 1998).…”

Genome-wide Analysis of AP-3–dependent Protein Transport in Yeast

“…Vacuole inheritance mutants can generate vacuoles de novo through an unknown mechanism (Weisman et al, 1987;Gomes De Mesquita et al, 1997). However, such vacuoles do not receive contents from the maternal vacuole that contains the active form of protease A responsible for maturation of ALP and CPS.…”