Characterisation of lead-induced stress molecular biomarkers in Medicago sativa plants

• We report the effects of Pb on physiological and transcriptional response in Alfalfa.
• Lipid peroxidation augmented remarkably, suggesting the onset of oxidative damages.
• A lack of PCs synthesis suggest a post-translational regulation.
• Alfalfa mitigate oxidative damage optimizing antioxidant enzymes and chaperones.

In this study, we investigated physiological changes and transcriptional responses in four-week-old hydroponically grown Medicago sativa seedlings exposed to (PbNO3)2 (0, 10 and 100 μM) for 2 and 7 days. Fresh weight and length were reduced in both shoots and roots after 7 days exposure. Lead accumulation was time and dose-dependent with stronger phytotoxic effects in roots than in shoots, with a relatively low amount of Pb translocated from roots to shoots. Lipid peroxidation augmented remarkably, suggesting the onset of oxidative damages, with increased glutathione reductase (GR), ascorbate peroxidase (APX) and superoxide dismutase (SOD) activities. This result was accompanied by a remarkable up-regulation of APX and SOD genes. In roots, the increase of SOD transcripts was concomitant to an enhanced SOD enzymatic activity in all Pb treatments. However, the 30-fold up-regulation occurred with a remarkable APX activity inhibition, which suggests that there might be post-transcriptional modifications able to regulate root APX. Root glutathione (GSH) and homoglutathione (hGSH) concentrations decreased in a dose dependent manner, while we could not detect the accumulation of phytochelatins (PC), albeit the related gene was up-regulated. The lack of PCs synthesis suggests a post-translational regulation of its enzymatic activity. Heat shock proteins (HSP70 and HSP17.7) were increased in alfalfa shoots, implying the triggering of cellular protection mechanism to cope with lead phytotoxicity. It is concluded that alfalfa plants mitigate the oxidative damage through induction of antioxidant enzymes, and the expression of chaperone proteins to alleviate Pb toxicity; metabolic changes that could be exploited as Pb-stress bioindicators.