Abstract
BackgroundMicroRNAs (miRNAs) are small, highly conserved, non-coding RNA that alter protein expression and regulate multiple intracellular processes, including those involved in the response to cellular stress. Alterations in miRNA expression may occur following exposure to several stress-inducing anticancer agents including ionizing radiation, etoposide, and hydrogen peroxide (H2O2).Methodology/Principal FindingsNormal human fibroblasts were exposed to radiation, H2O2, or etoposide at doses determined by clonogenic cell survival curves. Total RNA was extracted and miRNA expression was determined by microarray. Time course and radiation dose responses were determined using RT-PCR for individual miRNA species. Changes in miRNA expression were observed for 17 miRNA species following exposure to radiation, 23 after H2O2 treatment, and 45 after etoposide treatment. Substantial overlap between the miRNA expression changes between agents was observed suggesting a signature miRNA response to cell stress. Changes in the expression of selected miRNA species varied in response to radiation dose and time. Finally, production of reactive oxygen species (ROS) increased with increasing doses of radiation and pre-treatment with the thiol antioxidant cysteine decreased both ROS production and the miRNA response to radiation.ConclusionsThese results demonstrate a common miRNA expression signature in response to exogenous genotoxic agents including radiation, H2O2, and etoposide. Additionally, pre-treatment with cysteine prevented radiation-induced alterations in miRNA expression which suggests that miRNAs are responsive to oxidative stress. Taken together, these results imply that miRNAs play a role in cellular defense against exogenous stress and are involved in the generalized cellular response to genotoxic oxidative stress.
Highlights
Numerous adaptive mechanisms in cells alter gene expression in response to potentially lethal stressors
We show that alterations in miRNA expression due to ionizing radiation are produced in response to other agents that induce either DNA damage or oxidative stress
Results miRNA Microarray To determine if miRNA expression may be altered in response to exogenous agents that, at least in part, induce intracellular oxidative stress, 1522 primary human fibroblasts were exposed to doses of ionizing radiation, H2O2, or etoposide that had been previously determined to generate one log of cell kill by the clonogenic assay
Summary
Numerous adaptive mechanisms in cells alter gene expression in response to potentially lethal stressors. These mechanisms include the regulation of several fundamental cellular processes including cell cycle progression, gene transcription and translation, as well as epigenetic mechanisms such as DNA methylation or acetylation. It is likely that miRNAs regulate several genes involved in the cellular response to potentially lethal stressors. Changes in miRNA expression occur under conditions of hypoxia [3], and are localized to stress response elements in cells subjected to various stressors [4]. MicroRNAs (miRNAs) are small, highly conserved, non-coding RNA that alter protein expression and regulate multiple intracellular processes, including those involved in the response to cellular stress. Alterations in miRNA expression may occur following exposure to several stress-inducing anticancer agents including ionizing radiation, etoposide, and hydrogen peroxide (H2O2)
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