Understanding the molecular mechanism of ILV1 stress sensitivity in yeast by examination of differential patterns of gene expression between wild-type and ILV1 knockout strains of Saccharomyces cerevisiae
Barbosa, Jose; Giles, David K.
University of Tennessee at Chattanooga
Place of Publication
As arguably the most widely used industrial eukaryote, Saccharomyces cerevisiae has become one of the many model organisms used to study molecular genetics. A recent unpublished study at the University of Tennessee - Chattanooga showed that yeast cells that had the ILV1 gene knocked out had reduced survivability under stress conditions. This study then focused on the role of ILV1 in the stress response of cells by measuring the expression levels of 11 candidate genes that are all involved in some critical pathway in the yeast to see how gene expression was influenced as a result of the loss of the functional ILV1 gene. The cells were introduced to four high-stress conditions; salinity, oxidative, osmotic, and heat, followed by RNA isolation and a two-step qRT-PCR protocol to measure the relative mRNA within the cells after stress incubation. The results showed that the expression of each of the candidate genes was higher in the ILV1 knockout strain relative to that of the wildtype, BY4743 strain under normal physiological conditions. However, the overall expression of the candidate genes showed a decrease in the knockout strain under the four stress treatment conditions. A direct comparison in the expression levels of the knockout strain relative to the wildtype strain showed no significant differences in expression under the stress conditions. Although the final expression levels of the two strains were not significantly different, the results do not show the magnitude of the change in expression under the stress conditions between the strain. Taking into consideration the higher expression of the candidate genes under normal conditions, the ILV1 knockout strain was more affected under stress conditions and had a larger magnitude of change in expression than the wildtype strain. These findings are consistent with ILV1 being pleiotropic and playing a role in some type of stress response.
First, I would like to thank Dr. Margaret Kovach for serving as my thesis director and guiding me through this lengthy, detail-oriented process, as well as postponing her retirement to allow me to be her final undergraduate research student. I have learned so much over the past year and a half and none of it would have been possible without her guidance. Next, I would like to thank Dr. Jose Barbosa for not only providing the topic of this study but also for teaching me the incredible biochemical reactions that are occurring in this beautiful model organism. The lessons that I have learned from both of these advisors are ones that I will carry with me for the rest of my life. I also want to thank Dr. David Giles for agreeing to be on my DHON committee and allowing me to take up some of his time so that I may propose the work that I have done. Lastly, I would like to thank Dr. Ethan Carver for providing the funds for an additional batch of reverse transcriptase needed after we had discovered that we did not have enough RNA for qRT-PCR. Without his generosity, we may have not been able to finish this study.
B. S.; An honors thesis submitted to the faculty of the University of Tennessee at Chattanooga in partial fulfillment of the requirements of the degree of Bachelor of Science.
Saccharomyces cerevisiae--Effect of stress on; Gene expression; Yeast
Schuster, Emily, "Understanding the molecular mechanism of ILV1 stress sensitivity in yeast by examination of differential patterns of gene expression between wild-type and ILV1 knockout strains of Saccharomyces cerevisiae" (2022). Honors Theses.