Persistent oxidative stress and gene expression changes in radiation-induced genomic instability

Abstract

In addition to the damage caused by the initial exposure to ionizing radiation, it is now known that ionizing radiation induces delayed DNA damage in cells by a process known as radiation-induced genomic instability. Characterized using many different endpoints including increased mutation, frequency, increased level of reactive oxygen species (ROS), and decreased plating efficiency, radiation-induced genomic instability is best characterized in the GM10115 system by chromosomal instability. While the cellular and molecular mechanism of induced instability is not known, we are investigating the hypothesis that clones exhibiting instability display an altered pattern of gene expression. An 1152 gene microarray chip was used to uncover potential gene expression changes in two sets of two unstable clones in comparison to an irradiated but chromosomally stable clone. One candidate gene, Cu/Zn superoxide dismutase 1 (SOD1), was under-expressed in the unstable clones by an average of 3.66-fold by array analysis. Given the high levels of ROS observed in unstable clones, we performed a detailed analysis of SOD1 in four chromosomally unstable cell lines. Northern and western blotting assays could not confirm the microarray data, showing a decrease in expression of SOD1 in only one of four unstable lines, and increases in average protein level in all four unstable clones relative to control. Average SOD1 activity was also elevated in three of four unstable clones tested. These results indicate that while increased ROS levels are characteristic of many of our clones showing instability, this is not due to under-expression of SOD1 as suggested by DNA microarray analysis.