Comprehensive transcriptomic and genomic analysis of oxidative stress-resistant saccharomyces cerevisiae

Abstract

Oxidative stress occurs as the oxidant and antioxidant balance in an organism is disrupted in the direction of oxidants, and oxidants come to a position that can potentially cause damage. All aerobic organisms use oxygen for respiration and oxidation of nutrients for producing energy. For this reason, while oxygen is necessary for aerobic organisms to live, it is also harmful because of the reactive oxygen species (ROS) that emerge during these processes that are necessary for life. Lipid, protein and DNA molecules, which form the basis of life, can be damaged directly or indirectly due to oxidative stress caused by ROS. Aerobic organisms have developed various mechanisms to protect themselves from the harmful effects of oxidative stress. These mechanisms involve enzymatic and non-enzymatic (antioxidants) systems. In addition to these mechanisms, organisms use various repair mechanisms, especially to repair damaged DNA. Oxidative stress from ROS is biomedically important and widely researched, as it causes many neurodegenerative and autoimmune diseases, as well as cardiovascular diseases, cancer, and aging in mammals. The yeast Saccharomyces cerevisiae is used as a model organism, because of its beneficial characteristics. S. cerevisiae, which is a eukaryotic organism, has a short-life span. It is a single cell organism and can be found as haploid, diploid or polyploid in nature. Thus, S. cerevisiae is an important model organism for elucidating the processes in eukaryotic complex organisms. The aim of this study was to characterize an oxidative stress-resistant mutant S. cerevisiae strain at genetic and transcriptomic levels. The main idea, however, was to comprehensively examine the complex molecular infrastructure of oxidative stress resistance. In addition, as a physiological analysis, the cell wall properties of the mutant strain was also tested and information was obtained about the effect of stress on the cell wall structure. In this thesis study, transcriptomic analysis of the mutant S. cerevisiae was performed by comparing the transcriptomic profiles of the oxidative stress-resistant, evolved S. cerevisiae and the reference strain. Transcriptomic data of the strains were obtained by microarray analysis. As a genomic analysis, the entire genomes of the oxidative stress-resistant strain and the reference strain were sequenced and the differences in the genome were determined to find the mutations in the evolved srain that are related to oxidative stress resistance. In addition, cell wall analyses of the reference strain and the mutant strain were performed, using lyticase susceptibility test. As a result of transcriptomic analysis, it was observed that the expression level of 869 genes changed by at least 2-fold, 349 genes changed by at least 3-fold, 144 genes changed by at least 4-fold, and 67 genes changed by at least 5-fold in the oxidative stress-resistant mutant. Among these 869 genes that were differentially regulated by 2 times or more, 459 genes were upregulated and 410 genes were downregulated. The genes whose expression were decreased are generally related with ribosomal RNA, nuclear transport, organelle integration, tRNA cell cycle, mitosis and RNA polymerase. Expression levels of stress response genes, carbohydrates, lipid, protein, precursor metabolites and ion/metabolite transport-related genes were generally increased in the mutant strain. According to the ESR analysis results, a positive correlation was observed between the ESR-induced genes and the genes of the oxidative stress- resistant mutant with increased expression, according to the database. In addition, a positive correlation was observed between ESR-suppressed genes and genes of the mutant with decreased expression. The expression levels of two genes related to oxidative stress decreased and twenty three of them increased in the oxidative stress-resistant, evolved strain. The expression levels of six autophagy-related genes in the oxidative stress-resistant mutant decreased and 30 of them increased. According to whole genome sequencing results, 34 missense, 2 nonsense, 1 deletion and 12 silent mutations were found in the genes of the mutant strain, and 13 mutations were detected in chromosomal regions outside the coding regions. A nonsense mutation in the NRG1 gene, which is a transcriptomic regulator, results in the formation of a truncated protein. Further genomic and proteomic studies would be necessary to clarify the role of these genes and mutations in the oxidative stress resistance.