Metallothioneins 1 and 2 Have Distinct but Overlapping Expression Patterns in Arabidopsis 1 Margarita Garcı ´a-Herna ´ndez, Angus Murphy, and Lincoln Taiz* Biology Department, Sinsheimer Laboratories, University of California, Santa Cruz, California 95064 The spatial and temporal expression patterns of metallothionein (MT) isoforms MT1a and MT2a were investigated in vegetative and reproductive tissues of untreated and copper-treated Arabidopsis by in situ hybridization and by northern blotting. In control plants, MT1a mRNA was localized in leaf trichomes and in the vascular tissue in leaves, roots, flowers, and germinating embryos. In copper- treated plants, MT1a expression was also observed in the leaf mesophyll and in vascular tissue of developing siliques and seeds. In contrast, MT2a was expressed primarily in the trichomes of both untreated and copper-treated plants. In copper-treated plants, MT2a mRNA was also expressed in siliques. Northern-hybridization studies performed on developing seedlings and leaves showed tem- poral variations of MT1a gene expression but not of MT2a expres- sion. The possible implications of these findings for the cellular roles of MTs in plants are discussed. MTs are defined as low-M r , Cys-rich proteins that bind heavy metals. MTs are widely distributed in eukaryotic and prokaryotic organisms (for review, see Kagi, 1991; Robinson et al., 1993). In animals and fungi MTs have been shown to play a role in the detoxification of heavy metals, although their exact function is not completely understood. In plants a correlation has been observed between MT RNA levels and tolerance to heavy metals in different Arabidop- sis ecotypes (Murphy and Taiz, 1995a), suggesting a role in metal homeostasis in plants. In animals and yeast MT expression is regulated by metals (Robinson et al., 1993). In plants the effect of metals on the expression of MTs varies with the plant species, tissue, and MT type. In Mimulus guttatus (de Miranda, 1990), soybean (Kawashima, 1991), and barley (Okumura et al., 1991), MT mRNA levels were decreased by copper treatment, whereas in bean (Foley and Singh, 1994; Foley et al., 1997), wheat germ (Lane et al., 1987), and Nicotiana glutinosa (Choi et al., 1996), MT expression was not affected by metals. In Arabidopsis (Zhou and Goldsbrough, 1994, 1995; Murphy and Taiz, 1995a), wheat (Snowden and Gard- ner, 1993), pea (Evans et al., 1992), and rice (Hsieh et al., 1995), transcription of MTs was enhanced by certain metals only. As in animals (Robinson et al., 1993), a variety of other stimuli, including ABA, heat shock, cold shock, wounding, viral infection, senescence, salt stress, and Suc starvation, have been shown to influence expression of plant MTs (Buchanan-Wollaston, 1994, 1997; Foley and Singh, 1994; Hsieh et al., 1995; Murphy and Taiz, 1995a; Snowden et al., 1995; Choi et al., 1996; Foley et al., 1997). In Arabidopsis, three MT gene families have been iden- tified: MT1, MT2, and MT3 (Zhou and Goldsbrough, 1994; Murphy et al., 1997), homologs of which have been iden- tified in other species. The data available regarding the expression of MT genes from a variety of plant species indicate that each MT gene type exhibits characteristic temporal and tissue-specific expression patterns. Expres- sion of most MT1-like sequences has been detected primar- ily in roots (de Miranda et al., 1990; de Framond, 1991; Evans et al., 1992; Zhou and Goldsbrough, 1994; Hsieh et al., 1995; Hudspeth et al., 1996) and senescent leaves (Ka- washima et al., 1991; Buchanan-Wollaston, 1994, 1997; Hsieh et al., 1995; Foley et al., 1997). MT2-type transcripts have been detected primarily in leaves (Snowden and Gardner, 1993; Foley and Singh, 1994; Zhou and Golds- brough, 1994, 1995; Coupe et al., 1995; Choi et al., 1996) and roots of mature plants (Zhou, 1994; Snowden et al., 1995; Murphy, 1996). MT3-like mRNAs have been detected in leaves (Murphy, 1996; Bundithya and Goldsbrough, 1997), fruits (Ledger and Gardner, 1994), and developing em- bryos (Dong and Dunstan, 1996). In Arabidopsis each MT type appears to belong to a small gene family, the members of which appear to exhibit differential gene expression patterns. MT1 consists of three isoforms, MT1a, MT1b, and MT1c. MT1a is constitutively expressed in seedlings and is induced by copper in excised leaves, whereas MT1b seems to be a pseudogene. MT1c is expressed in young and mature roots and in mature leaves and is not affected by copper treatment. The MT2 gene family consists of MT2a and MT2b. They are both consti- tutively expressed in mature leaves, and only the MT2a gene is copper inducible in seedlings (Zhou and Golds- brough, 1994; Murphy and Taiz, 1995a; Zhou and Golds- brough, 1995; Murphy et al., 1997). Immunocytochemical studies have recently shown similar patterns of accumula- tion of the gene products (Murphy et al., 1997). All of these data suggest that each MT isoform may have specialized functions in different tissues. Some of the functions pro- posed for plant MTs include a role during development (Kawashima et al., 1992; Ledger and Gardner, 1994; Dong 1 This study was supported by a grant from the U.S. Department of Agriculture (no. 94-37100-0755) to L.T. and by a Research Sci- entist Training Postdoctoral Fellowship from the Ministry of Ed- ucation and Sciences of Spain to M.G.-H. * Corresponding author; e-mail taiz@biology.ucsc.edu; fax 1– 408 – 459 –3139. Abbreviations: MT, metallothionein; RT, reverse transcriptase. Plant Physiol. (1998) 118: 387–397 387 www.plantphysiol.org on July 30, 2016 - Published by www.plantphysiol.org Downloaded from Copyright © 1998 American Society of Plant Biologists. All rights reserved.