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BIOCHEMICAL PROCESSES AND MACROMOLECULAR STRUCTURES

Phytochelatin Synthases of the Model Legume Lotus japonicus. A Small Multigene Family with Differential Response to Cadmium and Alternatively Spliced Variants^1,[OA]

Departamento de Nutrición Vegetal, Estación Experimental de Aula Dei, Consejo Superior de Investigaciones Científicas, 50080 Zaragoza, Spain (J.R., M.R.C., L.N., J.L., C.P.-R., M.B.); and Kazusa DNA Research Institute, Kisarazu, Chiba 292–0818, Japan (S.S., S.T.)

The biosynthesis of phytochelatins and homophytochelatins has been studied in nodulated plants of the model legume Lotus (Lotus japonicus). In the first 6 to 24 h of treatment with cadmium (Cd), roots started to synthesize elevated amounts of both polypeptides, with a concomitant increase of glutathione and a decrease of homoglutathione, indicating the presence of active phytochelatin synthase (PCS) genes. Screening of transformation-competent artificial chromosome libraries allowed identification of a cluster of three genes, LjPCS1, LjPCS2, and LjPCS3, which were mapped at 69.0 cM on chromosome 1. The genes differ in exon-intron composition and responsiveness to Cd. Gene structures and phylogenetic analysis of the three protein products, LjPCS1-8R, LjPCS2-7N, and LjPCS3-7N, are consistent with two sequential gene duplication events during evolution of vascular plants. Two sites for alternative splicing in the primary transcripts were identified. One of them, involving intron 2 of the LjPCS2 gene, was confirmed by the finding of the two predicted mRNAs, encoding LjPCS2-7R in roots and LjPCS2-7N in nodules. The amino acid sequences of LjPCS2-7R (or LjPCS2-7N) and LjPCS3-7N share 90% identity, but have only 43% to 59% identity with respect to the typical PCS1 enzymes of Lotus and other plants. The unusual LjPCS2-7N and LjPCS3-7N proteins conferred Cd tolerance when expressed in yeast (Saccharomyces cerevisiae) cells, whereas the alternatively spliced form, LjPCS2-7R, differing only in a five-amino acid motif (GRKWK) did not. These results unveil complex regulatory mechanisms of PCS expression in legume tissues in response to heavy metals and probably to other developmental and environmental factors.

The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: Manuel Becana (becana{at}eead.csic.es).

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www.plantphysiol.org/cgi/doi/10.1104/pp.106.090894

^* Corresponding author; e-mail becana{at}eead.csic.es; fax 34–976–716145.

Received October 6, 2006; accepted December 26, 2006; published January 5, 2007.

This article has been cited by other articles:

				J. Ramos, L. Naya, M. Gay, J. Abian, and M. Becana Functional Characterization of an Unusual Phytochelatin Synthase, LjPCS3, of Lotus japonicus Plant Physiology, September 1, 2008; 148(1): 536 - 545. [Abstract] [Full Text] [PDF]

				M. C. Lopez-Martin, M. Becana, L. C. Romero, and C. Gotor Knocking Out Cytosolic Cysteine Synthesis Compromises the Antioxidant Capacity of the Cytosol to Maintain Discrete Concentrations of Hydrogen Peroxide in Arabidopsis Plant Physiology, June 1, 2008; 147(2): 562 - 572. [Abstract] [Full Text] [PDF]