Molecular cloning and characterization of a phytochelatin synthase gene, PvPCS1, from Pteris vittata L.

  • Ruibin Dong
  • Elide Formentin
  • Carmen Losseso
  • Francesco Carimi
  • Piero Benedetti
  • Mario Terzi
  • Fiorella Lo Schiavo
Environmental Biotechnology


Pteris vittata L. is a staggeringly efficient arsenic hyperaccumulator that has been shown to be capable of accumulating up to 23,000 μg arsenic g−1, and thus represents a species that may fully exploit the adaptive potential of plants to toxic metals. However, the molecular mechanisms of adaptation to toxic metal tolerance and hyperaccumulation remain unknown, and P. vittata genes related to metal detoxification have not yet been identified. Here, we report the isolation of a full-length cDNA sequence encoding a phytochelatin synthase (PCS) from P. vittata. The cDNA, designated PvPCS1, predicts a protein of 512 amino acids with a molecular weight of 56.9 kDa. Homology analysis of the PvPCS1 nucleotide sequence revealed that it has low identity with most known plant PCS genes except AyPCS1, and the homology is largely confined to two highly conserved regions near the 5′-end, where the similarity is as high as 85–95%. The amino acid sequence of PvPCS1 contains two Cys-Cys motifs and 12 single Cys, only 4 of which (Cys-56, Cys-90/91, and Cys-109) in the N-terminal half of the protein are conserved in other known PCS polypeptides. When expressed in Saccharomyces cerevisae, PvPCS1 mediated increased Cd tolerance. Cloning of the PCS gene from an arsenic hyperaccumulator may provide information that will help further our understanding of the genetic basis underlying toxic metal tolerance and hyperaccumulation.


Pteris vittata Phytochelatin Phytochelatin synthase Arsenic hyperaccumulation Heavy metal hypertolerance 



We thank Mr. Robert Tacchetto (Botanical Garden of Padua) for assisting in plant growth.


  1. 1.
    Altschul SF, Gish W, Miller EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410CrossRefPubMedGoogle Scholar
  2. 2.
    Clemens S, Kim EJ, Neumann D, Schroeder JI (1999) Tolerance to toxic metals by a gene family of phytochelatin synthases from plants and yeast. EMBO J 18:3325–3333CrossRefPubMedGoogle Scholar
  3. 3.
    Cobbett CS (2000) Phytochelatins and their roles in heavy metal detoxification. Plant Physiol 123:825–832CrossRefPubMedGoogle Scholar
  4. 4.
    Cobbett C (2003) Heavy metals and plants—model systems and hyperaccumulators. New Phytol 159:289–293CrossRefGoogle Scholar
  5. 5.
    Cobbett C, Goldsbrough P (2002) Phytochelatins and metallothioneins: roles in heavy metal detoxification and homeostasis. Annu Rev Plant Biol 53:159–182CrossRefPubMedGoogle Scholar
  6. 6.
    Ebbs S, Lau I, Ahner B, Kochian L (2002) Phytochelatin synthesis is not responsible for Cd tolerance in the Zn/Cd hyperaccumulator Thlaspi caerulescenes (J&C Presl). Planta 214:635–640CrossRefPubMedGoogle Scholar
  7. 7.
    Grill E, Winnacker EL, Zenk MH (1985) Phytochelatins: the principal heavy metal complexing peptides of higher plants. Science 230:674–676CrossRefGoogle Scholar
  8. 8.
    Grill E, Loeffler S, Winnacker EL, Zenk MH (1989) Phytochelatins, the heavy-metal-binding peptides of plants, are synthesized from glutathione by a specific glutamylcysteine dipeptidyl transpeptidase(phytochelatin synthase). Proc Natl Acad Sci USA 86:6838–6842CrossRefGoogle Scholar
  9. 9.
    Ha SB, Smith AP, Howden R, Dietrich WM, Bugg S, O’Connell MJ, Goldsborough PB, Cobbett CS (1999) Phytochelatin synthase genes from Arabidopsis and the yeast Schizosaccharomyces pombe. Plant Cell 11:1153–1163CrossRefPubMedGoogle Scholar
  10. 10.
    Hartley-Whitaker J, Ainsworth G, Vooijs R, Ten Bookum W, Schat H, Meharg AA (2001) Phytochelatins are involved in differential arsenate tolerance in Holcus lanatus. Plant Physiol 126:299–306PubMedCrossRefGoogle Scholar
  11. 11.
    Heiss S, Wachter A, Bogs J, Cobbett C, Rausch T (2003) Phytochelatin synthase (PCS) protein is induced in Brassica juncea leaves after prolonged Cd exposure. J Exp Bot 54:1833–1839PubMedCrossRefGoogle Scholar
  12. 12.
    Kojima Y, Binz PA, Kägi JH (1999) Nomenclature of metallothionein: proposal for a revision. In: Klaassen C (ed) Metallothionein IV. Birkhäuser, Basel, pp 7–13Google Scholar
  13. 13.
    Loeffler S, Hochberger A, Grill E, Winnacker EL, Zenk MH (1989) Termination of the phytochelatin synthase reaction through sequestration of heavy metals by the reaction product. FEBS Lett 258:42–46CrossRefGoogle Scholar
  14. 14.
    Ma LQ, Komar KM, Tu C, Zhang W, Cai Y, Kennelley ED (2001) A fern that hyperaccumulates arsenic. Nature 409:579CrossRefPubMedGoogle Scholar
  15. 15.
    Maier T, Yu C, Kullertz G, Clemens S (2003) Localization and functional characterization of metal-binding sites in phytochelatin synthases. Planta 218:300–308CrossRefPubMedGoogle Scholar
  16. 16.
    Maitani T, Kubota H, Sato K, Yamada T (1996) The composition of metals bound to class III metallothionein (phytochelatin and its desglycyl peptide) induced by various metals in root culture of Rubia tinctorum. Plant Physiol 110:1145–1150PubMedGoogle Scholar
  17. 17.
    Mizuno T, Toyoharu S, Kenji H, Keishi S, Akiyoshi T, Naoharu M, Hitoshi O (2003) Cloning and characterization of phytochelatin synthase from a nickel hyperaccumulator Thlaspi japonicum and its expression in yeast. Soil Sci Plant Nutr 49:285–290Google Scholar
  18. 18.
    Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  19. 19.
    Ortiz DF, Kreppel L, Speiser DM, Scheel G, McDonald G, Ow DW (1992) Heavy-metal tolerance in fission yeast requires an ATP-binding cassette-type vacuolar membrane transporter. EMBO J 11:3491–3499PubMedGoogle Scholar
  20. 20.
    Ortiz DF, Ruscitti T, McCue KF, Ow DW (1995) Transport of metal-binding peptides by HMT1, a fission yeast ABC-type vacuolar membrane protein. J Biol Chem 270:4721–4728CrossRefPubMedGoogle Scholar
  21. 21.
    Oven M, Page JE, Zenk MH, Kutchan TM (2002) Molecular characterization of the homo-phytochelatin synthase of soybean Glycine max: relation to phytochelatin synthase. J Biol Chem 277:4747–4754PubMedCrossRefGoogle Scholar
  22. 22.
    Rauser WE (1995) Phytochelatins and related peptides. Structure, biosynthesis and function. Plant Physiol 109:1141–1149CrossRefPubMedGoogle Scholar
  23. 23.
    Rea PA, Li ZS, Lu YP, Drozdowicz, YM, Martinoia E (1998) From vacuolar GS-X pumps to multispecific ABC transporters. Annu Rev Plant Physiol Plant Mol Biol 49:727–760PubMedCrossRefGoogle Scholar
  24. 24.
    Schat H, Llugany M, Vooijs R, Hartley-Whitaker J, Bleeker PM (2002) The role of phytochelatins in constitutive and adaptive heavy metal tolerances in hyperaccumulator and non-hyperaccumulator metallophytes. J Exp Bot 53:2381–2392CrossRefPubMedGoogle Scholar
  25. 25.
    Schmöger MEV, Oven M, Grill E (2000) Detoxification of arsenic by phytochelatins in plants. Plant Physiol 122:793–802CrossRefPubMedGoogle Scholar
  26. 26.
    Sneller FEC, Van Heerwaarden LM, Kraaijeveld-Smit FJL, Ten Bookum WM, Koevoets PLM, Schat H, Verkleij JAC (1999) Toxicity of arsenate in Silene vulgaris, accumulation and degradation of arsenate-induced phytochelatins. New Phytol 144:223–232CrossRefGoogle Scholar
  27. 27.
    Vatamaniuk OK, Mari S, Lu YP, Rea PA (1999) AtPCS1, a phytochelatin synthase from Arabidopsis: isolation and in vitro reconstitution. Proc Natl Acad Sci USA 96:7110–7115CrossRefPubMedGoogle Scholar
  28. 28.
    Vatamaniuk O, Mari S, Lu Y, Rea P (2000) Mechanism of heavy metal ion activation of phytochelatin (PC) synthase. Blocked thiols are sufficient for PC synthase-catalyzed transpeptidation of glutathione and related thiol peptides. J Biol Chem 275:31451–31459CrossRefPubMedGoogle Scholar
  29. 29.
    Vogeli-Lange R, Wagner GJ (1990) Subcellular localization of cadmium and cadmium-binding peptides in tobacco leaves: implication of a transport function for cadmium binding peptides. Plant Physiol 92:1086–1093CrossRefGoogle Scholar
  30. 30.
    Wang J, Zhao FJ, Meharg AA, Raab A, Feldmann J, McGrath SP (2002) Mechanisms of arsenic hyperaccumulation in Petris vittata. Uptake kinetics, interactions with phosphate, and arsenic speciation. Plant Physiol 130:1552–1561CrossRefPubMedGoogle Scholar
  31. 31.
    Zenk MH (1996) Heavy metal detoxification in higher plants—a review. Gene 179:21–30CrossRefPubMedGoogle Scholar
  32. 32.
    Zhao FJ, Wang JR, Barker JHA, Schat H, Bleeker PM, McGrath SP (2003) The role of phytochelatins in arsenic tolerance in the hyperaccumulator Pteris vittata. New Phytol 159:403–410CrossRefGoogle Scholar

Copyright information

© Society for Industrial Microbiology 2005

Authors and Affiliations

  • Ruibin Dong
    • 1
  • Elide Formentin
    • 1
  • Carmen Losseso
    • 1
  • Francesco Carimi
    • 1
  • Piero Benedetti
    • 1
  • Mario Terzi
    • 1
  • Fiorella Lo Schiavo
    • 1
  1. 1.Centro di Ricerca Interdipartimentale per le Biotechnologie InnovativeUniversita’ di PadovaPaduaItaly

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