Introduction
Oxidative stress can be defined as a decrease in the reducing capabilities of cellular redox agents. Cells maintain a reductive environment within themselves to function properly. Reactive oxygen species (ROS) and free radicals are produced naturally in organelles such as mitochondria and chloroplasts (Dietz 2008). Normally, these are removed by the cell. However, it is possible for ROS to overload the cell's ability to maintain a reductive environment. This "oxidative stress" can cause detrimental chemical changes within a cell. Oxidative stress causes cellular damage, and it has been linked to diseases in many organisms. In order for antioxidants to protect against ROS, NADPH must be present as a reducing agent for cellular redox reactions. About 60% of all NADPH in cells is derived from the pentose phosphate pathway (Kruger and Schaewen 2003). An enzyme, encoded by the gene ZWF1, catalyzes a major component of this pathway in yeast. A mutation in the ZWF gene leads to symptoms of oxidative stress such as hydrogen peroxide sensitivity (Slekar 2008). Over-expression of ZMS1 and ZMS2 genes relieves these symptoms of oxidative stress (Slekar 2008).
In this experiment, a microarray analysis was used to determine other yeast genes involved in cellular protection against oxidative stress. Isolated mRNA from ZMS1 and ZMS2 single knockout yeast were compared to mRNA isolated from wildtype yeast to determine changes in gene expression. Since ZMS1 and ZMS2 have been shown to play a role in regulating oxidative stress in yeast, this microarray analysis can give insight into other genes that play a role in this pathway. A microarray is a small glass or silicon chip with a chemical matrix to which DNA oglionucleotides can be covalently attached. This array can contain thousands of DNA sequences, and cDNAs with colored dyes can be used as probes to look for expression patterns. These dyes can be detected by a laser at the correct wavelength. By using microarray technology, we can examine differences in expression levels between ZMS2 knockout yeast and wildtype yeast for every gene in the yeast genome by comparative analysis.
Viewing these differences in expression between ZMS mutants and wildtype yeast cells can reveal why oxidative stress is relieved in the mutants. This can aid us in understanding and preventing the development of oxidative stress.
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