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Identification of transcriptome profiles for the DNA-damaging agents bleomycin and hydrogen peroxide in L5178Y mouse
Shawn D Seidel1, H Lynn Kan, William T Stott
1Toxicology & Environmental Research and Consulting, The Dow Chemical Company, Midland, Michigan 48674, USA. sseidel2@dow.com
Abstract:
It is believed that some aspects of genotoxicity are associated with changes in the transcription levels of certain genes, especially those involved in DNA repair and cell cycle control. Additionally, it is hypothesized that chemicals sharing a common mode of genotoxicity should exhibit similar changes in gene expression. We have evaluated these hypotheses by analyzing transcriptome profiles of mouse lymphoma L5178Y/TK(+/-) cells treated with bleomycin and hydrogen peroxide, two mutagens that produce genotoxicity by generating reactive free radicals. The cells were treated for 4 hr and RNA was isolated at the end of the treatment and after a 20 hr recovery. Transcriptome analyses were performed using the Clontech Mouse 1.2K cDNA microarray (1,185 genes) and hybridization with a (32)[P]-labeled probe. Of the genes examined, each mutagen altered the expression (1.5-fold or greater) of only two genes after the 4 hr treatment. In cells allowed to recover for 20 hr after treatment, bleomycin and hydrogen peroxide altered the expression of 8 and 5 genes, respectively. Many of the altered genes have some association with apoptosis. Of these genes, three (the genes encoding granzyme A, integrin beta 7, and 45 kDa calcium-binding protein precursor) were in common between chemical treatments. The expression of DNA repair and cell cycle controlling genes present on the array was not affected by the treatments. These results show that bleomycin and hydrogen peroxide both have unique and commonly regulated genes that have the potential to serve as biomarkers of exposure to agents causing DNA damage by free radical mechanisms.
Insights
Bleomycin and hydrogen peroxide exposure alters gene expression in mouse cells, revealing unique and common biomarkers for DNA damage from free radicals. These findings aid in identifying genotoxic agents.
Area of Science:
- Toxicology
- Molecular Biology
- Genetics
Background:
- Genotoxicity may involve altered gene transcription, particularly for DNA repair and cell cycle genes.
- Chemicals with similar genotoxic mechanisms are hypothesized to induce comparable gene expression changes.
Purpose of the Study:
- To investigate gene expression changes in response to bleomycin and hydrogen peroxide, two free radical-generating mutagens.
- To evaluate if shared genotoxic mechanisms correlate with similar transcriptome alterations.
- To identify potential biomarkers for DNA damage induced by free radical mechanisms.
Main Methods:
- Transcriptome profiling of mouse lymphoma L5178Y/TK(+/-) cells treated with bleomycin or hydrogen peroxide.
- Analysis of gene expression after 4-hour treatment and 20-hour recovery periods.
- Utilized Clontech Mouse 1.2K cDNA microarray (1,185 genes) and radioactive labeling for hybridization.
Main Results:
- Few genes (2 per mutagen) showed altered expression after 4 hours.
- After 20-hour recovery, bleomycin affected 8 genes and hydrogen peroxide affected 5 genes.
- Commonly affected genes included those related to apoptosis (granzyme A, integrin beta 7, 45 kDa calcium-binding protein precursor).
- No significant changes were observed in DNA repair or cell cycle control genes.
Conclusions:
- Bleomycin and hydrogen peroxide induce both unique and common gene expression changes.
- These identified genes may serve as biomarkers for exposure to free radical-induced DNA damage.
- Transcriptome analysis provides insights into the molecular responses to specific genotoxic agents.
