Browsing by Author "Ratnayake Mudiyanselage, Rasathma"

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    Examining the stress response in anhydrobiotic yeast (Saccharomyces cerevisiae)
    (2024-04-02) Ratnayake Mudiyanselage, Rasathma
    Yeast (Saccharomyces cerevisiae) is a unique model system for biological research due to the fact that it can be maintained in a dry desiccated state of anhydrobiosis for lengthy periods of time without the need for water or nutrients. However, there is very limited understanding of how anhydrobiosis impacts the natural stress response. The objective of this thesis was to study the thermal stress response in yeast pre- and post-desiccation. It was hypothesized that desiccation induces a stress response in yeast which makes them resistant to a thermal stress, and that as rehydrated yeast start to enter exponential growth, the stress response will be down regulated resulting in return to normal thermal sensitivity. To study this, two different strains of yeast were used: a wild type strain (YBS21-A) and a rad51 mutant strain (YBS29-1) that is defective in homologous recombinational DNA repair. Yeast samples were exposed to heat shocks pre- desiccation, as well at various timepoints post-rehydration. Survival was quantified at each timepoint following a heat challenge to identify shifts in thermal-sensitivity. In addition, transcriptional changes were quantified using RT-qPCR to identify which genes might be regulating the response. Immediately post rehydration, thermal survival to a potentially lethal heat challenge of 52oC for 10 minutes was high. Thermal tolerance then started to gradually decrease and survival decreased to up to three orders of magnitude by the 12 hour timepoint. Following this, heat tolerance began to increase back to almost 100% survival by 18 and 24 hours post rehydration. The trends in growth rate were opposite to that of survival. When cell growth rates were at their highest (exponential phase), thermal tolerance was low, whereas when growth rates were low (lag phase and stationary phase), thermal tolerance was high. The expression levels of multiple heat shock proteins were found to correlate with thermal survival, including hsp104, hsp78, hsp42 and hsp12, suggesting that they may be regulating thermal sensitivity. Interestingly, there was no difference in survival between the wild type and rad51 strains, suggesting that heat shock induced cell death is independent of DNA double strand break repair. Overall, this foundational knowledge of the stress response during anhydrobiosis and how it impacts thermal sensitivity is important for the use of desiccated yeast as a model organism for long term low maintenance biological research.