Molecular and physiological characterization of iron resistant Saccharomyces cerevisiae

Iron, the fourth most abundant element in earth crust, is essential for living organisms. At high concentrations, however, it is toxic. The electron transport potential between its two most common oxidation states (Fe3+ and Fe2+) makes iron crucial for all living organisms. Iron ions play critical roles in oxidative phosphorylation. Iron shows similar physical and chemical properties with other transition metals, like copper, zinc, nickel and cobalt. Transition metal ions have common transport systems or they can replace each other in some structures. Thus, it can be expected that iron resistance and resistance to other transition metals may have common features. The aim of this study was the molecular and physiological characterization of an iron-resistant Saccharomyces cerevisiae mutant that was previously obtained by evolutionary engineering, an inverse metabolic engineering strategy. For this purpose, the cross-resistance of the iron-resistant strain against other stress types (particularly other transition metal ions) were determined in detail. Growth physiological analyses were also performed, which revealed that the iron-resistant mutant produced high levels of glycerol and accumulated high amounts of trehalose. The yeast S. cerevisiae can survive toxic metals by either of the two major kinds of resistance mechanisms: sequestration or avoidance. To gain insight into the major iron resistance mechanism in the resistant strain, iron contents of the mutant and the reference strain were determined both in the presence and absence of iron stress, by using Flame Atomic Absorption Spectrophotometry (F-AAS). The results showed that the iron-resistant yeast strain had significantly higher iron level than the reference strain, already in the absence of any iron stress. When iron stress was present in the culture medium, however, the iron-resistant mutant did not take up as much iron ions as the reference strain did. These results imply that in the presence of iron stress, the iron-resistant mutant avoids iron ion uptake, to survive iron stress conditions. For molecular characterization of the iron-resistant S. cerevisiae strain, transcriptomic analyses were performed using quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) and microarray technology, both in the presence and absence of iron stress. Microarray analysis results showed that the iron-resistant S. cerevisiae strain had higher expression levels of the oxidative stress-related genes than the reference strain, even in the absence of any iron stress. According to the results obtained by the microarray analysis, lipid peroxidation assay and intracellular reactive oxygen species (ROS) levels determination assay were performed. The intracellular ROS level of the iron-resistant strain was found to be lower than that of the reference strain. Thus, the iron-resistant strain possibly expresses high levels of oxidative stress-related genes and produces antioxidant c which reduce intracellular ROS levels. By this means, it may survive high levels of iron stress.