Tolerance of Mannitol-Accumulating Transgenic Wheat to Water Stress and Salinity

Abstract

Previous work with model transgenic plants has demonstrated that cellular accumulation of mannitol can alleviate abiotic stress. Here, we show that ectopic expression of the mtlD gene for the biosynthesis of mannitol in wheat improves tolerance to water stress and salinity. Wheat (Triticum aestivum L. cv Bobwhite) was transformed with the mtlD gene of Escherichia coli. Tolerance to water stress and salinity was evaluated using calli and T 2 plants transformed with (ϩmtlD) or without (ϪmtlD) mtlD. Calli were exposed to Ϫ1.0 MPa of polyethylene glycol 8,000 or 100 mm NaCl. T 2 plants were stressed by withholding water or by adding 150 mm NaCl to the nutrient medium. Fresh weight of ϪmtlD calli was reduced by 40% in the presence of polyethylene glycol and 37% under NaCl stress. Growth of ϩmtlD calli was not affected by stress. In ϪmtlD plants, fresh weight, dry weight, plant height, and flag leaf length were reduced by 70%, 56%, 40%, and 45% compared with 40%, 8%, 18%, and 29%, respectively, in ϩmtlD plants. Salt stress reduced shoot fresh weight, dry weight, plant height, and flag leaf length by 77%, 73%, 25%, and 36% in ϪmtlD plants, respectively, compared with 50%, 30%, 12%, and 20% in ϩmtlD plants. However, the amount of mannitol accumulated in the callus and mature fifth leaf (1.7-3.7 mol g Ϫ1 fresh weight in the callus and 0.6-2.0 mol g Ϫ1 fresh weight in the leaf) was too small to protect against stress through osmotic adjustment. We conclude that the improved growth performance of mannitol-accumulating calli and mature leaves was due to other stress-protective functions of mannitol, although this study cannot rule out possible osmotic effects in growing regions of the plant.Water stress and salinity are major abiotic factors that limit crop productivity in drought-prone areas. One way of increasing productivity in stressful environments is to breed crops that are more tolerant to stress. However, success in breeding for tolerance has been limited because (a) tolerance to stress is controlled by many genes, and their simultaneous selection is difficult (Richards, 1996;Yeo, 1998;Flowers et al., 2000); (b) tremendous effort is required to eliminate undesirable genes that are also incorporated during breeding (Richards, 1996); and (c) there is a lack of efficient selection procedures particularly under field conditions (Ribaut et al., 1997). Genetic engineering offers an alternative approach for developing tolerant crops. Unlike classical breeding, genetic engineering is a faster and more precise means of achieving improved tolerance (Cushman and Bohnert, 2000) because it avoids the transfer of unwanted chromosomal regions. Moreover, through genetic engineering, multiple genes can be assembled and simultaneously introduced to the crop of interest. There are many functional targets for engineering tolerance to water stress and salinity, one of them being accumulation of osmoprotectants (Rathinasabapathi, 2000).The osmolyte mannitol is normally synthesized in numerous plant species, but not in wheat (Triticum aes...

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