Functional Characterization of NRAMP3 and NRAMP4 from the Metal Hyperaccumulator Thlaspi caerulescens

• Published in: The New phytologist Vol. 181; no. 3; pp. 637 - 650
• Main Authors: Oomen, Ronald J. F. J., Wu, Jian, Lelièvre, Françoise, Blanchet, Sandrine, Richaud, Pierre, Barbier-Brygoo, Hélène, Aarts, Mark G. M., Thomine, Sébastien
• Format: Journal Article
• Language: English
• Published: Oxford, UK Blackwell Publishing 01.02.2009; Blackwell Publishing Ltd; Wiley
• Subjects: Arabidopsis; Arabidopsis - drug effects; Arabidopsis - metabolism; arabidopsis-thaliana; Biological Transport; Biological Transport - drug effects; Cadmium; cadmium (Cd); Cadmium - toxicity; Cloning, Molecular; Complementary DNA; DNA, Complementary; DNA, Complementary - genetics; DNA, Plant; DNA, Plant - metabolism; expression differences; gene encodes; Gene Expression Regulation, Plant; Gene Expression Regulation, Plant - drug effects; Genes; Genetic Complementation Test; Genome, Plant; Genome, Plant - genetics; Green Fluorescent Proteins; Green Fluorescent Proteins - metabolism; heavy-metal; Homeostasis; hyperaccumulator; Hyperaccumulators; Hypersensitivity; Intracellular Membranes; Intracellular Membranes - drug effects; Intracellular Membranes - metabolism; Iron; iron transport; Life Sciences; metal transport; Metals; Metals - metabolism; Molecular Sequence Data; Mutation; Mutation - genetics; NRAMP; Plant Proteins; Plant Proteins - genetics; Plant Proteins - metabolism; Plants; protein; Recombinant Fusion Proteins; Recombinant Fusion Proteins - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - cytology; Saccharomyces cerevisiae - drug effects; Thlaspi; Thlaspi - drug effects; Thlaspi - genetics; Thlaspi - metabolism; Thlaspi caerulescens; transporter genes; Vacuoles; Vacuoles - drug effects; Vacuoles - metabolism; Vegetal Biology; Yeasts; Zinc; zinc (Zn); Zinc - toxicity; zinc transporter; NRAMP; hyperaccumulator; zinc (Zn); metal transport; arabidopsis-thaliana; Thlaspi caerulescens; cadmium (Cd)
• ISSN: 0028-646X; 1469-8137
• DOI: 10.1111/j.1469-8137.2008.02694.x

The ability of metal hyperaccumulating plants to tolerate and accumulate heavy metals results from adaptations of metal homeostasis. NRAMP metal transporters were found to be highly expressed in some hyperaccumulating plant species. Here, we identified TcNRAMP3 and TcNRAMP4, the closest homologues to AtNRAMP3 and AtNRAMP4 in Thlaspi caerulescens and characterized them by expression analysis, confocal imaging and heterologous expression in yeast and Arabidopsis thaliana. TcNRAMP3 and TcNRAMP4 are expressed at higher levels than their A. thaliana homologues. When expressed in yeast TcNRAMP3 and TcNRAMP4 transport the same metals as their respective A. thaliana orthologues: iron (Fe), manganese (Mn) and cadmium (Cd) but not zinc (Zn) for NRAMP3; Fe, Mn, Cd and Zn for NRAMP4. They also localize at the vacuolar membrane in A. thaliana protoplasts. Inactivation of AtNRAMP3 and AtNRAMP4 in A. thaliana results in strong Cd and Zn hypersensitivity, which is fully rescued by TcNRAMP3 or TcNRAMP4 expression. However, metal tolerance conferred by TcNRAMP expression in nramp3nramp4 mutant does not exceed that of wild-type A. thaliana. Our data indicate that the difference between TcNRAMP3 and TcNRAMP4 and their A. thaliana orthologues does not lie in a different protein function, but probably resides in a different expression level or expression pattern.