Functional characterization of metallothionein gene family in Arabidopsis

Woei-Jiun Guo, Purdue University

Abstract

In Arabidopsis thaliana, metallothioneins (MTs) are encoded by a seven member gene family. To examine the tissue-specific expression of MT genes, GUS reporter gene activity driven by promoters of MT1a, MT2a, MT2b and MT3 was analyzed in transgenic plants. MT1a and MT2b are expressed in the phloem of all organs and are copper (Cu) - inducible; MT2a and MT3, in contrast, are expressed predominantly in mesophyll cells and are also induced by Cu in young leaves and in root apecies. Expression of MT4 genes is restricted to seeds. When exressed in metal-sensitive yeast mutants, Arabidopsis MTs that are expressed primarily in vegetative tissues (MTI, MT2 and MT3) restored Cu and Cd tolerance, whereas MT4 proteins were more effective for Zn tolerance and accumulation. MT-GFP proteins localized in the cytosol of yeast and plant cells. These results indicate that all seven MTs are functional cytoplasmic metal chelators, although with specific binding properties. To examine the roles of MTs in plants, I have identified Arabidopsis mutants that are deficient in MT1a, MT1c, MT2a, MT2b and MT3 via T-DNA insertions or RNAi interference. These mutations, either singly or in combination, have no effect on growth under normal conditions, suggesting plant MTs are not essential for normal plant development. However, when plants were exposed to excess Cu, lack of MT1a reduced Cu content in roots without affecting Cu accumulation in shoots. In contrast, Cu accumulation in mt2a and mt2b was not significantly affected and no additive effect was seen in plants that lacked MT1a, MT2a and MT2b. However, plants deficient in MT2a and MT2b, but not MT1a, were sensitive to Cu and Cd and hyper-sensitive to these metals in the absence of phytochelatins. These results indicate that individual MTs have specific contributions to metal homeostasis. MT1a, which is highly induced in phloem, plays a prominent role in Cu distribution in roots. MT2a, MT2b and phytochelatins function cooperatively in metal tolerance, especially for Cd detoxification.

Degree

Ph.D.

Advisors

Goldsbrough, Purdue University.

Subject Area

Botany

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