Genotype differences towards lead chloride harmful action

Abstract

The aim of the study was to throw more light on the PbCl2 mode of action (MoA) depending on the genotype by the application of three model organisms and microbiological, biochemical, and molecular approaches. Three model systems – Chlamydomonas reinhardtii strain 137C – wild type (WT), Saccharomyces cerevisiae strain D7ts1, and Pisum sativum L. cultivar Ran1 and two experimental schemes – short- and long-term treatments were used. C. reinhardtii and S. cerevisiae cell suspensions (1×106 cells/ml) at the end of the exponential and the beginning of a stationary phase of growth were treated with various PbCl2 concentrations (0.45–3.6 mM) for 2 hours. Lower PbCl2 concentrations (0.03–0.22 mM) were also tested on C. reinhardtii 137C. Short-term treatment for up to 2 days with PbCl2 concentrations in the range of 0.45–3.6 mM and long-term treatment for up to 10 days with concentrations in the range of 0.45–2.7 mM was performed on P. sativum L. seeds and plants, respectively. Long-term treatment with a PbCl2 concentration of 3.6 mM was not tested because of the very strong toxic effect (plant death). The following endpoints were used – for C. reinhardtii: cell survival, “visible” mutations, DNA double-strand breaks (DSBs), malondialdehyde (MDA), intracellular peroxides (H2O2), and photosynthetic pigments; for S. cerevisiae – cell survival, gene conversion, reverse mutation, mitotic crossing-over, DSBs, superoxide anions, MDA and glutathione (GSH); P. sativum L. – germination and root length (short-term treatment), pro-oxidative markers – MDA, H2O2 and photosynthetic pigments (long-term treatment). Genotype differences between C. reinhardtii (0.047 mM) and S. cerevisiae (1.66 mM) were observed by two endpoints: concentrations inducing 50% lethality (LD50) and DSB induction. By contrast, no mutagenic effect was found for both unicellular test models. A slight toxic capacity of PbCl2, measured as inhibition of Pisum sativum L. seed germination and around 20% root length reduction was revealed after the treatment with concentrations equal to or higher than 1.8 mM. The variety of stress responses between the two plant test models was demonstrated by comparing MDA and H2O2. A dose-dependent increase in H2O2 levels and a minor increase of MDA levels (around 9–15%) were measured when C. reinhardtii cells were treated with concentrations in the range of LD20–LD80 (0.03–0.11 mM). Analyzing the kinetics of MDA and H2O2 in pea leaves, the most pronounced effect of concentration was shown for 2.7 mM. A decrease in the photosynthetic pigments was detected in the two experimental designs – short-term on C. reinhardtii and long-term on P. sativum treatments. The pro-oxidative potential was also proven in S. cerevisiae based on increased levels of MDA and superoxide anions and decreased GSH. New information is gained that PbCl2 can affect the DNA molecule and photosynthetic pigments via induction of oxidative stress. Our study revealed that the magnitude of stress response towards PbCl2 is genotype-specific. Our finding that Chlamydomonas reinhardtii is a sensitive test system towards PbCl2 contributes to good strategies for revealing very low levels of contaminants present chronically in main environmental matrices. This is the first report, as far as we know, affirming that PbCl2 can induce DSBs in Chlamydomonas reinhardtii and Saccharomyces cerevisiae.