Figure 1.
Phytochelatin-2 is present at higher levels than phytochelatin-3 in earthworms, and both are arsenic-responsive in laboratory exposures.
Figure 2.
Glycine and methionine are the NMR-detectable metabolites most responsive to arsenic.
Significance of rank correlations of all metabolites with soil arsenic concentrations (Spearman’s μ).
Figure 3.
Earthworm phytochelatin synthase sequences compared to phytochelatin synthase sequences from other species.
A: alignment of Lumbricus rubellus phytochelatin synthase sequences (conserved N-terminal region only, full sequence shown in Figure S1) against key species with validated PCS activity (Arabidopsis thaliana PCS1 and PCS2, Eisenia fetida, L. rubellus PCS1a and PCS1b, Caenorhabditis elegans, and Schizosaccharomyces pombe). Conserved residues are shown with white text on black background; the conserved Cys-His-Asp catalytic triad is shown with yellow on blue background. B: Phylogenetic tree of phytochelatin synthase genes. The evolutionary history was inferred by using the Maximum Likelihood method based on the Whelan and Goldman model [48]. The tree with the highest log likelihood (-3323.4018) is shown. The percentage of trees in which the associated taxa clustered together is shown next to the branches. Initial tree(s) for the heuristic search were obtained by applying the Neighbor-Joining method to a matrix of pairwise distances estimated using a JTT model. A discrete Gamma distribution was used to model evolutionary rate differences among sites (5 categories (+G, parameter = 1.9794)). The tree is drawn to scale, with branch lengths measured in the number of substitutions per site. The analysis involved 19 amino acid sequences. All positions containing gaps and missing data were eliminated. There were a total of 126 positions in the final dataset. Evolutionary analyses were conducted in MEGA5 [49]. At – A. thaliana. Bj – Brassica juncea. Nt – Nicotiana tabacum. Df – Dictyostelium fasciculatum. Cg – Crassostrea gigas. Sj yFS275 – Schizosaccharomyces japonicum. Sp – S. pombe. Cb – C. briggsae. Ce1a – C. elegans. Lr1a and Lr1b – Lumbricus rubellus PCs 1a and 1b respectively. Cs – Clonorchis sinensis. Sj – Schistosoma japonicum. Sm – Schistosoma mansoni. Ci – Ciona intestinalis. Sp1 and Sp2 – Strongylocentrotus purpuratus.
Figure 4.
phytochelatin synthase isoforms were not arsenic responsive at the concentrations and exposure duration tested. Filled bars: pcs-1a. Empty bars: pcs-1b. Error bars represent 95% confidence intervals (n = 5).
Figure 5.
Phytochelatins are elevated in worms from contaminated field sites (blue) compared to worms from relatively pristine control sites (red).
A: phytochelatin-2; B: phytochelatin-3. The boxes display the median, and first and third quartile boundaries, and the whiskers represent 95% confidence intervals. DGC1: Devon Great Consols ‘control’ site, in effect a second arsenic-contaminated site. DGC2: Devon Great Consols contaminated site. SHP1: Shipham contaminated site. SHP2: Shipham control site. AHT: Alice Holt. DRA: Drayton. SND: Snowdon.