Cyclic peptides-cyclotides biosynthesized by Viola species strengthen the tolerance to heavy metals acting as a defense system

The Viola genus is rich in species tolerant to heavy metals. Long term studies showed that both metallicolous (MET - occurring on polluted areas) and non-metallicolous (NMET - growing on non-polluted soils) species tolerate high concentrations of heavy metals (Zn and Pb) at the level of the whole plant and isolated cells in suspension. Metal tolerance depends on many factors, among which the secondary metabolites produced by plants play an important role. The species of Violaceae family produce cyclic peptides-cyclotides whose primary function is defense against insect pests. The question arises whether cyclotides could play a role in plant protection against abiotic factors, including heavy metals?. The Viola species/genotypes, representing different tolerance level to heavy metals (MET, NMET) were selected as a source material for the study. The first step was to detect the distribution of heavy metals (Zn, Pb) in cell organelles using cell suspension culture and transmission electron microscopy with X-ray microanalysis. Pb or Zn in V. tricolor NMET cells were deposited in vacuoles and cell wall, small particles were also found in cytoplasm and nucleus (Sychta et al. ۲۰۱۸). Cyclotides biosynthesis was determined in plants collected from MET and NMET soils and in plants (NMET and MET genotype) growing under in vitro conditions. The LC-MS technique showed that V. tricolor plants collected from MET soils produced more cyclotides than plants from NMET sites. The plants (NMET, MET genotype) growing under in vitro conditions (control, free from heavy metals) did not differ in the production of cyclotides. It suggest that the presence of heavy metals stimulate cyclotide production. The cyclotide biosynthesis pattern was also determined in suspended cells of NMET and MET Viola species treated with ۲۰۰ and ۲۰۰۰ μM of zinc or lead for ۲۴ and ۷۲ h, using MALDI-MS technique. Results showed that most of cyclotides were overproduced in the MET genotypes, contrary to NMET genotypes with not increased content of cyclotides after metal treatment. The biosynthesis of cyclotides after treatment with heavy metals was increased, especially in treatments with high concentration of Zn and low doses of Pb.Conclusion: The increased biosynthesis of cyclotides after exposure to heavy metals, in plants from MET sites and the deposition of cyclotides and heavy metals in vacuoles indicates that these peptides may strengthen heavy metal tolerance in violets. Also the presence of heavy metal binding sites in cyclotide chemical structure suggests that these peptides could have an important share in heavy metal binding and detoxification.

کلیدواژه ها:

Heavy metal tolerance ، cyclotides ، metallophytes ، dotoxification.