All living cells appear to be capable of exiting the normal cell cycle (proliferating state) and entering an alternative (resting) state termed quiescence. Most eukaryotic cells, whether they exist as single-celled or multicellular organisms, spend the majority of their natural lives in a quiescent state. Beyond contributing to a more thorough understanding of the life cycle of cells, studying quiescence has other potentially significant implications; a deeper grasp of the conserved mechanisms underlying entry into, survival in, and exit from quiescence in eukaryotes may aid the development of immunosuppressants and anti-fungal therapies and is also likely to provide significant insights into diverse processes as aging and growth control.
One of the simplest eukaryotic organisms, and a popular model organism for biological research is the budding yeast Saccharomyces cerevisiae (S. cerevisiae). The organism’s response to its transition from growth to quiescence is complex and polymorphic. Although several polymorphisms that influence sporulation, which is a response to nutrient depletion that allows a single diploid cell to give rise to four stress-resistant haploid spores have been identified, those that promote cellular quiescence are much less well understood. Dr. Linda Breeden and members of her laboratory in the Basic Sciences division have been studying this growth to quiescence transition using a combination of genetics, genomics and biochemistry, and reported their findings in a recent issue of the journal Molecular Biology of the Cell.
Dr. Breeden describes their work: “Diploid yeast can either sporulate or enter a quiescent cellular state. These fate choices show broad natural variations that are very reproducible. In this paper we show that the highly conserved NDR/Lats and MAP kinase signaling cascades promote quiescence and defects therein lead to cell death. The transition in and out of quiescence is key to survival for most organisms. With this unicellular model eukaryote, we can follow these transitions without intervention and identify both conserved and fungi-specific steps, both of which could be medically important.”
Using wild and laboratory strains of S. cerevisiae, the authors showed that the ability to enter quiescence is a highly variable but reproducible quantitative trait, by assaying strains as haploids and diploids for their ability to enter a stress tolerant and long-lived quiescent state compared to their ability to sporulate. In lab strains, sporulation is triggered by glucose and nitrogen deprivation; however, several wild strains have lost the requirement for nitrogen starvation and 100% will sporulate. Disrupting sporulation by deleting both copies of the IME1 gene enabled one of these diploids to enter quiescence, suggesting that there is a decision-making hierarchy that favors sporulation in some strains. W303 is a lab yeast strain where the pathway to quiescence is largely restricted to haploids due to the presence of the truncated and defective ssd1-2 gene. Ssd1 is an RNA-binding protein that is highly pleiotropic and interacts with more than 800 genes. It is regulated by the conserved NDR/Lats (Large tumor suppressor) kinase that is also involved in cell cycle control and development in humans. They found that the function of Ssd1 is more critical in diploids than in haploids for entry into quiescence. This is likely due to its role in cell wall remodeling; there is evidence that cell wall fortification is a key step in the transition to quiescence, and loss of Ssd1 function confers sensitivity to cell wall disrupting agents. The authors show in this work that Ssd1 is indeed crucial for the formation and longevity of quiescent cells in W303 diploids. They also report that the Cell Wall Integrity pathway is required for surviving the entry into quiescence when Ssd1 function is compromised.
Dr. Breeden explains the significance of this work: “We know a lot about the cell division process in rapidly growing yeast, and we know that this process is fundamentally conserved from yeast to humans. There is much less known about how the cell cycle is stably but reversibly arrested in quiescence, in spite of the fact that most cells spend most of their time in the quiescent state. After all, a cancer cell is a cell that fails to enter quiescence or exits from that state when it shouldn’t. A stem cell has to remain in a protected quiescent state until it is signaled to re-enter the cell cycle for tissue remodeling and renewal.”
Indeed, the tendency to either sporulate or enter quiescence displayed by these yeast strains suggest there has been selective pressure to limit pathway choice to promote longevity in different environmental conditions. As for what comes next for the Breeden lab: “Just testing the roles of known cell division regulators in the transitions in and out of quiescence could occupy decades of research, but with yeast we can use genetics to select for genes that promote or prevent the transition to quiescence or increase life span in that non-dividing state. Our ‘Fountain of Youth’ project is a cross between a short-lived and a long-lived yeast strain where we select survivors that can recover from a year in the quiescent state. One of the things we have found is mutant versions of Ssd1 that increase or reduce longevity. Ssd1 is a fungi-specific protein, and as such, is a candidate for anti-fungal therapy,” said Dr. Breeden.
Miles S, Li LH, Melville Z, Breeden LL. 2019. Ssd1 and the Cell Wall Integrity Pathway promote entry, maintenance and recovery from the quiescent state in budding yeast. Mol Biol Cell, May 29. [Epub ahead of print]
This work was supported by the National Institutes of Health.