Isolation of quiescent and nonquiescent cells from yeast stationary-phase cultures.
Quiescence is the most common and, arguably, most poorly understood cell cycle state. This is in part because pure populations of quiescent cells are typically difficult to isolate. We report the isolation and characterization of quiescent and nonquiescent cells from stationary-phase (SP) yeast cultures by density-gradient centrifugation. Quiescent cells are dense, unbudded daughter cells formed after glucose exhaustion. They synchronously reenter the mitotic cell cycle, suggesting that they are in a G(0) state. Nonquiescent cells are less dense, heterogeneous, and composed of replicatively older, asynchronous cells that rapidly lose the ability to reproduce. Microscopic and flow cytometric analysis revealed that nonquiescent cells accumulate more reactive oxygen species than quiescent cells, and over 21 d, about half exhibit signs of apoptosis and necrosis. The ability to isolate both quiescent and nonquiescent yeast cells from SP cultures provides a novel, tractable experimental system for studies of quiescence, chronological and replicative aging, apoptosis, and the cell cycle.
Allen
Chris
C
Department of Biology, Health Sciences Center, University of New Mexico, Albuquerque, NM 87131, USA.
Büttner
Sabrina
S
Aragon
Anthony D
AD
Thomas
Jason A
JA
Meirelles
Osorio
O
Jaetao
Jason E
JE
Benn
Don
D
Ruby
Stephanie W
SW
Veenhuis
Marten
M
Madeo
Frank
F
Werner-Washburne
Margaret
M
D017209:Apoptosis / Q000502:physiology; D002453:Cell Cycle / Q000502:physiology; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient / Q000379:methods; D005434:Flow Cytometry; D005947:Glucose / Q000737:chemistry; D008853:Microscopy / Q000379:methods; D008938:Mitosis; D017382:Reactive Oxygen Species / Q000378:metabolism; D016192:Resting Phase, Cell Cycle / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
D017382:Reactive Oxygen Species; D005947:Glucose