ORIGINAL ARTICLE Stephen Ebbs á Ingar Lau á Beth Ahner á Leon Kochian Phytochelatin synthesis is not responsible for Cd tolerance in the Zn/Cd hyperaccumulator Thlaspi caerulescens J. & C. Presl) Received: 6 February 2001 /Accepted: 3 May 2001 / Published online: 29 September 2001 Ó Springer-Verlag 2001 Abstract Thlaspi caerulescens J. & C. Presl, ``Prayon'') is a heavy-metal hyperaccumulator that accumulates Zn and Cd to high concentrations 40,000 and 4,000 mg kg DW ±1 , respectively) without phytotoxicity. The mecha- nism of Cd tolerance has not been characterized but reportedly involves vacuolar sequestration. The role of phytochelatins PCs) in metal tolerance in T. caerules- cens and the related non-accumulator T. arvense was examined. Although PCs were produced by both species in response to Cd, these peptides do not appear to be involved in metal tolerance in the hyperaccumulator. Leaf and root PC levels for both species showed a sim- ilar positive correlation with tissue Cd, but total PC levels in the hyperaccumulator were generally lower, despite correspondingly higher metal concentrations. The lack of a role for PCs in the hyperaccumulator's response to metal stress suggests that other mechanisms are responsible Cd tolerance. The lower level of leaf PCs in T. caerulescens also implies that Cd in the shoot is sequestered in a compartment or form that does not elicit a PC response. Keywords Cadmium á Hyperaccumulator á Metal tolerance á Phytochelatin á Thlaspi Cd tolerance) Abbreviations GSH: glutathione á PC: phytochelatin Introduction Thlaspi caerulescens J. & C. Presl) is a well-known hyperaccumulator of heavy metals. While Zn hyper- accumulation and tolerance have received considerable attention VaÂzquez et al. 1992, 1994; Baker et al. 1994; Lasat et al. 1996, 1998; KuÈpper et al. 1999; Salt et al. 1999), the comparable mechanisms for Cd have not. Zinc tolerance is achieved by sequestration in the leaf epidermal and mesophyll vacuoles VaÂzquez et al. 1992, 1994; Lasat et al. 1998; Salt et al. 1999). Since phytochelatin PC)-Cd complexes are also sequestered in cell vacuoles Salt and Rauser 1995), Cd tolerance in T. caerulescens may be due to increased PC syn- thesis. However, PCs do not contribute to the Cd tolerance observed in metal-tolerant populations of Silene vulgaris L. bladder campion) De Knecht et al. 1992, 1994, 1995) and other species. Since these species are taxonomically and morphologically dis- similar, their tolerance mechanisms may dier. The objective of this study was to determine if PCs play a role in Cd hyperaccumulation and tolerance in T. caerulescens. Materials and methods Hydroponic culture methods Seeds of Thlaspi caerulescens J. & C. Presl, ecotype. Prayon) and the non-accumulator Thlaspi arvense Crucifer Genetics Cooperative, Madison, Wis., USA) were grown hydroponically in aerated 2-l pots of nutrient solution Ebbs et al. 1997). Much of the research into metal transport and tolerance in T. caerulescens has utilized the Prayon ecotype Lasat et al. 1996, 1998; Lombi et al. 2000; Luo et al. 2000; McGrath et al. 2000; Pence et al. 2000), and Cd accumulation has been documented. Plants were grown for 30±40 days to equivalent biomass, with the nutrient solution changed weekly. Plants were treated for 4 days with Cd 2+ as CdSO 4 ) at the following concentrations: 0, 1, 2, 5, 15, 25, 40, 50 lM. Roots were rinsed at harvest to desorb residual treatment solution. Planta 2002) 214: 635±640 DOI 10.1007/s004250100650 S. Ebbs &) Department of Plant Biology, Southern Illinois University Carbondale, Carbondale, IL 62901, USA E-mail: sebbs@plant.siu.edu Fax: +1-618-4533441 I. Lau á B. Ahner Department of Agricultural and Biological Engineering, Cornell University, Ithaca, NY 14853, USA L. Kochian U.S. Plant, Soil, and Nutrition Laboratory, USDA-ARS, Cornell University, Ithaca, NY 14853, USA