"We have only begun to explore life on Earth." "The vast
majority of the cells in your body are not your own; they belong to bacterial
and other microorganismic species."
Edward O. Wilson
"and most of these are quiescent." -WW lab
Most of the cells you know are quiescent! Like cultures of other microorganisms, cultures of the yeast Saccharomyces cerevisiae responds to starvation by arresting growth and entering stationary phase. We define the arrested cultures as being in stationary phase, and the state of cells that are able to withstand long periods without added nutrients as being quiescent. Because most microorganisms exist under conditions of nutrient limitation, the ability to survive starvation is critical for survival. Molecular analyses have identified changes in transcription, translation, and protein modification in stationary-phase cells. At the level of translation, the pattern of newly synthesized proteins in stationary-phase cells is surprisingly similar to those synthesized during exponential growth. When limited for different nutrients, yeast strains may not enter stationary phase but opt for pathways such as pseudohyphal growth. If nutrient limitation continues, the endpoint is likely to be a quiescent cell.
The importance of the quiescent state is that although most cells on the planet are quiescent, the requirements for entry, survival, and exit are not well understood. This state is extremely important for pathogenesis (Tuberculosis and Cryptococcus infections), bioterrorism (germination of Anthrax spores), and other areas of biology, including ecology (un-cultivatable microorganisms). We have, through genomic analysis, identified novel mutants that are essential for survival in the quiescent state and look forward to identifying novel genes required for re-entry into the cell cycle.
We are currently using the yeast GFP-fusion library (4159 strains each with the Green Fluorescent Protein gene inserted 3' of the ORF) to identify proteins that accumulate in quiescent or non-quiescent cells. We have screened the almost 25,000 strains in exponential and stationary phases and are following up with these studies microscopically and using a variety of flow cells and microchannel devices. With the GFP proteins we have identified, we are finally able to identify the trajectory for the formation of quiescent and non-quisecent cells.
We also are doing a highthroughput screen of GFP strains with and without the TOR inhibitor rapamycin. TOR is a highly conserved signal transduction pathway. We have identified novel proteins that are induced and repressed by rapamycin and have used these to screen 300,000 chemicals in the NIH library to identify novel TOR inhibitors and activators. This work has been done in collaboration with the UNM Flow Cytometry Facility.
We are also developing new technologies aimed at increasing the throughput of proteomics assays and providing novel information about protein expression.
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