Biology Metal accumulating plants Mechanisms of metal hyperaccumulation in plants

1. Biology • Metal accumulating plants • Mechanisms of metal hyperaccumulation in plants • Mechanisms of metal resistance: Phytochelatins and metallothioneins • Molecular mechanisms of ion transport in plant cells

2. Metal accumulating plants

3. Bioavailability of metals to hyperaccumulators

7. Observation of Zn accumulation in plants was first recorded in 1865 by F. Risse (German scientist?). The plant, Thlaspi alperstre var. calaminare grown in zinc-rich soil in a area between Germany and Belgium. The leave tissue of the plant contained Zn exceeding 10,000 mg Zn/kg (1% dry matter), or 10% Zn in the ash. • Observations of unusual accumulation of other metals have been made only during the twentieth century. E.g. Pb, 1920s, Se 1930s, Ni, 1940s, Co and Cu, 1960s, Cd and Mn, 1970s.

8. Example: In 1930s, Se was found to be responsible for “alkali disease” in range animals in South Dakota. Plants, in the genus of Astragalus, are capable of accumulating up to 0.6% Se in dry shoot biomass.

9. At least 45 plant families are known to contain metal accumulating species and 397 metal accumulating taxa have been identified

10. Hyperaccumulators of Ni Family species location Max. Conc (mg/kg) Asteraceae Berkheya coddii South Africa 11,600 Pentacalia (10 species) Cuba 16,600 Brassicaceae Bornmuellera (6 taxa) Greece 17,600 Peltaria emarginata Greece 34,400 Streptanthus polygaloides USA(CA) 14,800 Rubiaceae Psychotria costivenia Cuba 38,530 P. vanhermanii Cuba 35,720

11. Hyperaccumulators of Zn, Cd and Pb Family species location Max. Conc (mg/kg) Zn Cd Pb Brassicaceae Thlaspi caerulescens W&Centr Europe 43,710 2,130 2,740 Caryophyllaceae Minuartia verna Yugoslavia; UK 11,400 20,000 Dichapetalaceae Dichapetalum gelonioides Sumatra; Mindanao; Sabah 30,000

12. Hyperaccumulators of Cu and Co (from Democratic Republic of Congo) Family species Max. Conc (mg/kg) Cu Co Convolvulaceae Ipomoea alpina 12,300 Lamiaceae Aeollanthus subacaulis var. linearis 13,700 4,300 Haumaniastrum katangense 9,222 2,241 H. robertii 2,070 10,232

13. Hyperaccumulators of Mn (from New Caledonia) Family species Max. Conc (mg/kg) Mn Celastracear Maytenus bureaviana 33,750 M. sebertiana 22,500 Proteaceae Macadamia angustifolia 11,590 M. neurophylla 55,200

14. Hyperaccumulators of Se (from New Caledonia) Family species Location Max. Conc (mg/kg) Asteraceae Haplopappus condensata Midwest USA 9,120 Brassicaceae Stanleya pinnata Midwest USA 1,190 S. bipinnata Midwest USA 2,380 Lecythidaceae Lecythis ollaria Venezuela 18,200 Leguminosae Astragalus bisulcatus Midwest USA 8,840 A. racemosus Midwest USA 14,920

15. Mechanisms of metal hyperaccumulation in plants

16. Definition of an essential element 1. If plant cannot complete its life cycle in the absence of the element 2. It forms part of any molecule of constituent of the plant that is itself essential in the plant

17. 16 elements are believed to be essential for plant growth. These are: C, H, O, N, P, K, S, Ca, Mg, B, Cl, Cu, Fe, Mn, Mo, Zn In addition to the 16 elements essential for plants, higher animals require sodium, iodine, cobalt, selenium, nickel, silicon, chromium, tin, vanadium, and fluorine, but not boron. (including boron, total 25 26 for animals)

18. Salisbury and Ross, 1985

19. Metal toxicity • Genetic variation • May be absorbed only to a limited extent, avoidance than true tolerance • Accumulate in roots with little transport to shoots • Both roots and shoots contain much higher amounts of such elements than nontolerant species could live with • Mechanism of true tolerance have not been understood • Suggested mechanisms: • Formation of stable nontoxic chelates • Storage of elements in vacuoles

20. Mechanisms of metal hyperaccumulation • Rhizosphere Interactions • Root uptake • Root-to-Shoot metal translocation • Metal sequestration and complexation

21. Rhizosphere Interactions • Hyperaccumulator species are able to accumulate higher metal concentrations in their shoots than surrounding nonaccumulator plants even from soils containing nonphytoxic background levels of metals. • Possibly due to enhance ability to solubilize metals within the rhizosphere of the hyperaccumulator. • By • The release of specific metal-chelating compounds into the rhizosphere by plant roots • modification of the rhizosphere pH or redox potential by plant roots.

22. Root uptake • Hyperaccumulation does not appear to be driven by the enhance affinity of root uptake systems for the hyperaccumulated metal, but increased rates of toot uptake. • Possiblly • enhanced expression of metal transporter. E.g. Roots of the zinc hyperaccumulator T. caerulescens appear to contain more zinc transporters per gram fresh weight than the nonaccumulator T. arvense.

23. Some plants demonstrate metal selectivity Metal selectivity could be due to metal transport across the root plasma membrane during either metal uptake into the symplast or metal export into the xylem

24. Root-to-Shoot metal translocation • Limited evidence indicated that rates of metal translocation from root to shoot are similar in hyperaccumulators and related nonaccumulator species. • Possibly, hyperaccumulators may lack the ability to restrict metal movement into the shoot. • Shoot:root ratio of metal concentrations are above unity in hyperaccumulators of Ni, Zn, or Co, suggesting an efficient root-to-shoot translocation system for the hyperaccumulated metals.

25. Metal sequestration and complexation • Metal hyperaccumulators tend to accumulate metals in epidermal and subepidermal tissues, including leaf trichomes. • Nickle and zinc are predominantly localized in vacuoles • Metal toxicity is reduced by complexing with high affinity ligands or organic acids • Evidence: many Ni and Zn hyperaccumulators accumulate high concentrations of organic acids in their leaves.

26. A young dicot root

27. A typical root

28. Basis tissue pattern in a mature root

29. Summary Evidence suggested that several mechanisms of hyperacumulation have been involved for one metal Hyperaccumulation may require several processes: Increased root uptake as well as reduced root accumulation, sequestration at cellular level as well as the tissue level, and, most importantly metal tolerance.

30. Mechanisms of metal resistance: • Phytochelatins and metallothioneins

31. Wenzel et al., 1999

32. Plants have adaptive mechanisms to respond to both nutrient deficiencies and toxicity. Metal tolerance is possibly related to: • Metal binding to cell walls • Metal tolerance of the membrane • Reduced membrane transport • Active efflux of metals form the cells-plants • Metal-tolerant enzymes • Compartmentation • Chelation of the metal by organic or inorganic ligands • Precipitation of metal compounds with low solubility

33. There are two major heavy metal-binding compounds in plant cells: The phytochelatin peptides (PCs) and metallothioneins (MTs).

34. MTs: • Low molecular weight (<10 kDa) • Large fraction of cystein residues • High metal content with coordination of metal ions in metal–thiolate clusters

35. MTs and PCs have been classified into three classes: • Class I: MTs from mammals and other organisms with a highly conserved arrangement of cystein residues. • Class II: all other MT proteins. • Class III: cystein-rich, metal-binding peptides that are not produced by translation of a mRNA on ribosome and therefore includes PCs.

36. Phytochelatin • Composed of only three amino acids: glutamate, cysteine, and glycine • Not coded directly by genes but likely to be products of biosynthetic pathways, presumably using GSH (glutathione) as a substrate • Have been identified in a wide variety of plant species, algae, fungal species and marine diatoms.

38. Synthesis of PCs can be induced by a wide range of metal ions, including Cd, Ni, Cu, Zn, Ag, Sn, Sb, Te, W, Au, Hg, Pb, and Bi • Cd was the most effective inducer. • Exposure of Cd in the range of 1-100 mM, induction can be detected within hours of exposure.

39. Mechanism seems a lot more complex than simply chelating the metal ion. • The metal ion activate PC synthase, be chelated by the PCs • Be transported to the vacuole and possibly form a more complex aggregation in the vacuole with , for example, sulfide or organic acids

40. The only MT proteins that have been purified from plants are the wheat Ec protein and a number of MTs from Arabidopsis. There are striking similarity in the cystein-rich domains which may have been duplicated within a single MT gene.

41. Presence of heavy metals may induce the expression of MT genes. Type I, II and III MTs are more expressed in root than in leaves; while type IV MTs are expressed in developing seeds MTs are also expressed in senescing tissues because they are possibly involved in metal ion transport in this process.

42. Summary • A role for PCs in the detoxification of some heavy metals, particularly Cd, is clearly established. • MTs are likely involved in metal metabolism in plants. However, their role in phytoremediation is highly speculative at this time.

43. Molecular mechanisms of ion transport in plant cells

44. Apoplastic--The interconnecting walls and the water-filled xylem elements are considered as a single system Symplastic--the rest of the plant, the “living” part, including the cytoplasm of all the cells in the plant. The cytoplasm of adjoining cells is connected through plasmodesmata in the cell walls.

46. Movement of ions via the apoplastic pathway can occur through walls of cortex cells until restricted by the impermeable Casparian strips of endodermal cells. • Regardless of the pathway across the root, ions transported to the shoot must somehow get into dead conducting cells of the xylem.

48. Transport across the plant plasma membrane is driven by an electrochemical gradient of protons generated by plasma membrane H+-ATPase Many genes encoding transporters have been identified and cloned.

49. These mustard plants carry a gene that helps them soak up heavy metals (Philip Rea, University of Pennsylvania)

50. An experimental treatment wetland at the Department of Energy's Savannah River Site tests the ability of native aquatic plants to clean up the acidic, metal contaminated runoff from a coal pile (University of Georgia Savannah River Ecology Laboratory)