Initial results in the development of a reporter cell line for toxicology studies at gene expression level: activation of the electrophile-responsive element by copper and methyl parathion.

Abstract

Induction of many genes encoding detoxifying enzymes and antioxidant proteins is mediated through a common mechanism, which is controlled by electrophile-responsive elements (EpRE) within the regulatory region of those genes. Copper and methyl parathion are environmental pollutants known to induce the expression of EpRE-mediated genes. In order to evaluate the molecular response triggered by these pollutants, a stable cell line was produced, which carries a transgene comprised of the green fluorescent protein (GFP) reporter gene under transcriptional control of the mouse glutathione-S-transferase (gst1) electrophile-responsive element fused to the mouse metallothionein (mt1) minimal promoter. The rat HTC hepatoma cells were transfected with the EpREmt-GFP construct and successfully selected with G418 antibiotic. EpREmt-GFP HTC cells were treated with 0.002 mg L(-1), 0.02 mg L(-1), 0.2 mg L(-1) and 2 mg L(-1) copper sulfate and 0.001 mg L(-1), 0.01 mg L(-1), 0.1 mg L(-1) and 1 mg L(-1) methyl parathion for 48 h. GFP expression was directly quantified in living cells using a microplate fluorimeter. GFP expression was significantly increased in higher concentrations of both pollutants, showing a 1.80- and 2.58-fold induction of GFP at 2mg copper L(-1) and 1mg methyl parathion L(-1), respectively (p<0.01). The results obtained in the present study demonstrate that the EpREmt-GFP HTC cell line can be an interesting model for further development for the study of the cellular response to aquatic pollutants as well as a new tool for environmental monitoring at the molecular level.

• Authors: Daniela Volcan Almeida, Gilma Santos Trindade, Laura A Geracitano, Daniela Martí Barros, José Maria Monserrat, Luis Fernando Marins
• Journal: Marine environmental research (Mar Environ Res) Vol. 66 Issue 1 Pg. 158-60 (Jul 2008) ISSN: 0141-1136 [Print] England
• PMID: 18533248 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Water Pollutants, Chemical
• Green Fluorescent Proteins
• Methyl Parathion
• Copper

Topics

• Animals
• Cell Line, Tumor
• Copper (toxicity)
• Ecotoxicology (methods)
• Gene Expression Profiling
• Gene Expression Regulation (drug effects)
• Genes, Reporter (genetics)
• Green Fluorescent Proteins (genetics)
• Methyl Parathion (toxicity)
• Mice
• Rats
• Reproducibility of Results
• Toxicity Tests (methods)
• Transgenes (genetics)
• Water Pollutants, Chemical (toxicity)