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A rapid and efficient method to study the function of crop plant transporters in Arabidopsis

Xiangfeng Wang, Fudi Zhong, Cheuk Hang Woo, Yansong Miao, Michael A. Grusak, Xiaobo Zhang, Jumin Tu, Yum Shing Wong, Liwen Jiang
Protoplasma 2016 pp. -
Arabidopsis, cell culture, confocal microscopy, food crops, gain-of-function mutation, genes, heterologous gene expression, humans, iron, mutants, nutrient deficiencies, rapid methods, rice, seeds, space and time, staple foods, transgenic plants, transporters, vacuoles, yeasts, zinc
Iron (Fe) is an essential micronutrient for humans. Fe deficiency disease is wide-spread and has lead to extensive studies on the mechanisms of Fe uptake and storage, especially in staple food crops such as rice. However, studies of functionally related genes in rice and other crops are often time and space demanding. Here, we demonstrate that transgenic Arabidopsis suspension culture cells and Arabidopsis plants can be used as an efficient expression system for gain-of-function study of selected transporters, using Fe transporters as a proof-of-principle. The vacuolar membrane transporters OsVIT1 and OsVIT2 have been described to be important for iron sequestration, and disruption of these two genes leads to Fe accumulation in rice seeds. In this study, we have taken advantage of the fluorescence-tagged protein GFP-OsVIT1, which functionally complements the ccc1 yeast mutant, to generate transgenic Arabidopsis suspension cell lines and plants. GFP-OsVIT1 was shown to localize on the vacuolar membrane using confocal microscopy and immunogold EM. More importantly, the Fe concentration, as well as the concentration of Zn, in the transgenic cell lines and plants were significantly increased compared to that in the WT. Taken together, our study shows that the heterologous expression of rice vacuolar membrane transporter OsVIT1 in Arabidopsis system is functional and effectively enhances iron accumulation, indicating a rapid and efficient approach for studying other putative transporters of crop plants in this system.