Characteristics of accumulation and tolerance of cadmium (Cd) in green manure crops were investigated to identify Cd-accumulating crops and to clarify the mechanisms involved in Cd accumulation and tolerance. Seedlings of eight crop species were treated with Cd (1 mg l(-1) or 5 mg l(-1)) in the growing medium for 4 d. Cd concentration in leaves of Avena strigosa Schreb. cv. New-oat, Crotalaria juncea L. and Tagetes erecta L. cv. African-tall was greater than values used to define Cd-hyperaccumulation (>100 mg Cd kg(-1) DW). However, in leaves of T. erecta, lipid peroxidation level increased significantly, and the activities of superoxide dismutase, ascorbate peroxidase, glutathione reductase, and catalase were depressed by both Cd treatments. By contrast, A. strigosa and C. juncea exhibited high Cd tolerance. Avena strigosa leaves showed higher activities of antioxidative enzymes such as superoxide dismutase and ascorbate peroxidase than those of other species tested. Crotalaria juncea showed higher amounts of total soluble phenolics which, in leaves, were doubled by 5 mg l(-1) Cd treatment. When two Cd-tolerant accumulators (A. strigosa and C. juncea) and the non-accumulator (C. spectabilis) were treated with lower Cd concentrations for 4 weeks, A. strigosa and C. juncea exhibited superior Cd accumulation in the shoots with greater biomass production compared with C. spectabilis. These results indicate that A. strigosa and C. juncea possess the greater potential for Cd accumulation and tolerance than common crops.

a b s t r a c tIn order to clarify the physiological role of the merE gene of transposon Tn21, a pE4 plasmid that contained the merR gene of plasmid pMR26 from Pseudomonas strain K-62, and the merE gene of Tn21 from the Shigella flexneri plasmid NR1 (R100) was constructed. Bacteria with plasmid pE4 (merR-o/p-merE) were more hypersensitive to CH 3 Hg(I) and Hg(II), and took up significantly more CH 3 Hg(I) and Hg(II), than the isogenic strain. The MerE protein encoded by pE4 was localized in the membrane cell fraction, but not in the soluble fraction. Based on these experimental results, we suggest for the first time that the merE gene is a broad mercury transporter mediating the transport of both CH 3 Hg(I) and Hg(II) across the bacterial membrane.

The characteristics of bacteria take up mercury into cells via membrane potential-dependent sequencedivergent members of the mercuric ion (Mer) superfamily, i.e., a periplasmic mercuric ion scavenging protein (MerP) and one or more inner membrane-spanning proteins (MerC, MerE, MerF, and MerT), which transport mercuric ions into the cytoplasm, have been applied in engineering of bioreactor used for mercurial bioremediation. We engineered bacteria to express MerC, MerE, MerF, or MerT with or without MerP to clarify their individual role and potential in transport of mercurial. By immunoblot analysis using specific polyclonal antibody, the proteins encoded by merC, merE, merF, merT or merP, were certainly expressed and identified in the membrane fraction. Bacteria expressing MerC, MerE, MerF or MerT in the absence of MerP transported significantly more C 6 H 5 Hg(I) and Hg(II) across bacterial membrane than their isogenic strain. In vivo expression of MerP in the presence of all the transporters did not cause apparent difference to the C 6 H 5 Hg(I) transport, but gives an apparently higher Hg(II) transport than that did by MerE, MerF or MerT but not by MerC. Among the four transporters studied, MerC showed more potential to transport Hg(II) across bacterial membrane than MerE, MerF and MerT. Together these findings, we demonstrated for the first time that in addition to MerE and MerT, MerF and MerC are broad-spectrum mercury transporters that mediate both Hg(II) and phenylmercury transport into cells. Our results suggested that MerC is the most efficient tool for designing mercurial bioremediation systems, because MerC is sufficient for mercurial transport into cells.

Phytochelatin (PC) synthesis has been well demonstrated as a major metal tolerance mechanism in Arabidopsis thaliana, whereas its contribution to long-distance element transport especially in monocots remains elusive. Using rice as a cereal model, we examined physiological roles of Oryza sativa phytochelatin synthase 1 (OsPCS1) in the distribution and detoxification of arsenic (As) and cadmium (Cd), two toxic elements associated with major food safety concerns. First, we isolated four different transcript variants of OsPCS1 as well as one from OsPCS2. Quantitative real-time reverse transcription–PCR (RT-PCR) of each OsPCS transcript in rice seedlings suggested that expression of OsPCS1full, the longest OsPCS1 variant, was most abundant, followed by OsPCS2. Heterologous expression of OsPCS variants in PCS-deficient mutants of Schizosaccharomyces pombe and A. thaliana suggested that OsPCS1full possessed PCS activity in response to As(III) and Cd while the activity of other PCS variants was very low. To address physiological functions in toxic element tolerance and accumulation, two independent OsPCS1 mutant rice lines (a T-DNA and a Tos17 insertion line) were identified. The OsPCS1 mutants exhibited increased sensitivity to As(III) and Cd in hydroponic experiments, showing the importance of OsPCS1-dependent PC synthesis for rice As(III) and Cd tolerance. Elemental analyses of rice plants grown in soil with environmentally relevant As and Cd concentrations showed increased As accumulation and decreased Cd accumulation in grains of the T-DNA line. The Tos17 mutant also exhibited the reduced Cd accumulation phenotype. These contrasting effects on As and Cd distribution to grains suggest the existence of at least partially distinct PC-dependent pathways for As and Cd.

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