Real-time luminescence monitoring of cell-cycle and respiratory oscillations in yeast.
The use of luciferase reporters has become a precise, noninvasive, high-throughput method for real-time monitoring of promoter activity in living cells, especially for rhythmic biological processes such as circadian rhythms. We developed a destabilized firefly luciferase as a reporter for rhythmic promoter activity in both the cell division and respiratory cycles of the budding yeast Saccharomyces cerevisiae in which real-time luminescence reporters have not been previously applied. The continuous output of light from luciferase reporters allowed us to explore the relationship between the cell division cycle and the yeast respiratory oscillation, including the observation of responses to chemicals that cause phase shifting of the respiratory oscillations. Destabilized firefly luciferase is a good reporter of cell cycle position in synchronized or partially synchronized yeast cultures, in both batch and continuous cultures. In addition, the oxygen dependence of luciferase can be used under certain conditions as a genetically encodable oxygen monitor. Finally, we use this reporter to show that there is a direct correlation between premature induction of cell division and phase resetting of the respiratory oscillation under the continuous culture conditions tested.
Robertson
J Brian
JB
Department of Biological Sciences, Vanderbilt University, Nashville, TN 37240, USA.
Stowers
Chris C
CC
Boczko
Erik
E
Johnson
Carl Hirschie
CH
D000079:Acetaldehyde / Q000494:pharmacology; D000332:Aerobiosis / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D000431:Ethanol / Q000494:pharmacology; D017930:Genes, Reporter; D008156:Luciferases / Q000378:metabolism; D008163:Luminescent Measurements / Q000379:methods*; D010100:Oxygen / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D013997:Time Factors
D000431:Ethanol; D008156:Luciferases; D000079:Acetaldehyde; D010100:Oxygen