Analysis of DNA damage and repair of plant extracts and phytochemicals, using yeasts as model organism

Abstract

In this work the model organism Saccharomyces cerevisiae was used to characterise the biological activity and the mechanism of action of plant extracts and phytochemicals. One of the goals was to use mutant strains affected in oxidative stress response and DNA repair in order to uncover the molecular targets of plant extracts and phytochemicals. DNA damage was assessed with the comet assay [1,2], using “comet tail length” as a measure of DNA damage, which displayed a dose-response relationship with different DNA-damaging agents. Typical experiments involved incubation of yeast cells with plant extracts or phytochemicals before, during or after (for DNA repair kinetics) the oxidative shock with hydrogen peroxide. The results obtained with the comet assay showed that DNA damage was significantly decreased upon treatment with plant extracts and several phytochemicals in a dose-dependent manner (p< 0.001). In addition, DNA repair kinetics was significantly improved in cells incubated with an extract of leafs from Ginkgo biloba (GBE) [3]. However, in the mutant strain affected in CDC9, encoding a DNA ligase involved in the mechanisms of nucleotide excision repair and base excision repair, oxidative DNA damage repair kinetics was unchanged with GBE, suggesting that the activity of this extract involves one of these mechanisms, or both. As expected, GBE treatment improved survival of yeast cells when challenged with oxidative shock by H2O2. In addition, intracellular oxidation, measured by flow cytometry with the redox-sensitive fluorochrome dichlorofluorescein diacetate, was considerably decreased upon pre-treatment with GBE [3]. The use of simple eukaryotic organisms and the use of genetically manipulated strains can provide tools for the analysis of the mechanism of action of the biological activity of phytochemicals and plant extracts.