Long-distance root-to-shoot transport of phytochelatins and cadmium in Arabidopsis

Abstract

Phytochelatin synthases (PCS) mediate cellular heavy-metal resistance in plants, fungi, and worms. However, phytochelatins (PCs) are generally considered to function as intracellular heavymetal detoxification mechanisms, and whether long-distance transport of PCs occurs during heavy-metal detoxification remains unknown. Here, wheat TaPCS1 cDNA expression was either targeted to Arabidopsis roots with the Arabidopsis alcohol dehydrogenase (Adh) promoter (Adh:: TaPCS1͞cad1 C admium is a widespread nonessential toxic heavy metal, released into the biosphere mostly by modern industry (1). Phytochelatins (PCs) play an essential role in heavy-metal detoxification in plants and fungi (2, 3). PCs chelate heavy metals and then PC-metal complexes are translocated across the tonoplast and sequestered in vacuoles (4, 5), thus decreasing the heavy-metal content in the cytosol of yeast and plant cells. PCs are synthesized from glutathione by the enzyme PC synthase (PCS) (6-9). However, PC research has focused mainly on PC transport into vacuoles (4, 5), and it remains unknown whether long-distance transport of PCs or PCS occurs in plants.Cd 2ϩ is taken up by plants, and plants have been proposed to provide an efficient system for heavy-metal removal from soils (10, 11). Soil composition affects Cd 2ϩ sensitivity. For example, silicon in soil reduces heavy-metal sensitivity (12, 13). Normally, Cd 2ϩ concentrations in roots are at least 10 times greater than those in shoots (14). However, for efficient phytoextraction from soils, heavy metals must be translocated into aerial tissues for later harvest. Translocation of Cd 2ϩ from roots to shoots has been studied in diverse plant species (15-18) and has been proposed to occur via the xylem of Indian mustard in a PCindependent manner (16). The molecular mechanisms for rootto-shoot Cd 2ϩ transport remain largely unknown, and PCs are predicted not to undergo long-distance transport but to enhance vacuolar heavy-metal sequestration in the cells in which they are produced.The cad1-3 mutant is a recessive, loss-of-function mutation in the Arabidopsis AtPCS1 gene (7) (9) by PCR. The TaPCS1 PCR fragment was subcloned into pBluescript II SK(ϩ) vector. A 3ϫ myc tag DNA sequence was constructed by PCR recovery from a plasmid containing c-myc and then subcloned into the pGEM-T Easy vector (Promega). The subcloned myc fragment was recovered with SpeI and SacI enzymes and then fused to the 3Ј end of the TaPCS1 ORF in the pBluescript II SK(ϩ) vector at the SpeI site. All PCR products were confirmed by sequencing (Retrogen, La Jolla, CA). The fusion sequence was then digested with BamHI and SacI and subcloned into the binary expression vector pBI121, either with the cauliflower mosaic virus 35S or the Arabidopsis alcohol dehydrogenase (Adh) promoter (20). Both constructs were transformed into the PC-deficient Arabidopsis mutant cad1-3 by direct Agrobacterium tumefaciens-mediated transformation using the floral dip technique (21). T2 seeds with 3:1 segregation on kanamycin plates were used for hom...

H+‐pyrophosphatase IbVP1 promotes efficient iron use in sweet potato [Ipomoea batatas(L.) Lam.]

“…Sweet potato seedlings grown in hydroponics were washed as described (Gong et al ., 2003) before sampling root and leaf tissues. The control plant tissues and plant tissues treated for Fe deficiency were digested in concentrated nitric acid for 5–7 days at room temperature, and the samples were boiled and completely digested.…”

Reduced translocation of cadmium from roots is associated with increased production of phytochelatins and their precursors

“…Previously, the role of PCs was thought to be limited to the intracellular detoxification mechanism by shuttling Cd 2+ -PC n complexes into the vacuole. However, recent studies on Brassica napus (Mendoza-Cózatl et al, 2008) and 22 Arabidopsis thaliana (Gong et al, 2003;Chen et al, 2006) showed that PCs could also play a major role in long-distance transport of Cd 2+ through xylem and phloem. Mendoza-Cózatl et al (2008) found that the concentration of PCs was 50 times higher in the phloem sap compared to the xylem sap and concluded that phloem was more active in transporting Cd 2+ from the source (older leaves) to the sink tissues (root, branches, younger leaves).…”

Examining the Specific Contributions of Individual Arabidopsis Metallothioneins to Copper Distribution and Metal Tolerance

“…PCs may also have a role in the transport of Cu in the xylem or phloem tissues. It has already been shown that PCs can translocate from roots to shoots and vice versa, and that PCs mediate root-to-shoot transport of Cd (Gong et al, 2003;Chen et al, 2006).…”

“…In particular, the cad1-3 mutant was shown to be PC deficient and hypersensitive to Cd toxicity due to a mutation in the AtPCS1 gene (Howden and Cobbett, 1992;Howden et al, 1995;Ha et al, 1999). Recent evidence also shows that PCs participate in transport of Cd from roots to shoots as a mechanism to alleviate Cd accumulation in Arabidopsis root cells (Gong et al, 2003).…”