To investigate the potential role of a polyol, mannitol, in oxidative stress protection, a bacterial mannitol-1-phosphate dehydrogenase gene was targeted to chloroplasts by the addition of an amino-terminal transit peptide. Transgenic tobacco (Nicotiana tabacum) lines accumulate mannitol at concentrations ranging from 2.5 to 7 mumol/g fresh weight. Line BS1-31 accumulated approximately 100 mM mannitol in chloroplasts and was identical to the wild type in phenotype and photosynthetic performance. The presence of mannitol in chloroplasts resulted in an increased resistance to methyl viologen (MV)-induced oxidative stress, documented by the increased retention of chlorophyll in transgenic leaf tissue following MV treatment. In the presence of MV, isolated mesophyll cells of BS1-31 exhibited higher CO2 fixation than the wild type. When the hydroxyl radical probe dimethyl sulfoxide was introduced into cells, the initial formation rate of methane sulfinic acid was significantly lower in cells containing mannitol in the chloroplast compartment than in wild-type cells, indicating an increased hydroxyl radical-scavenging capacity in BS1-31 tobacco. We suggest that the chloroplast location of mannitol can supplement endogenous radical-scavenging mechanisms and reduce oxidative damage of cells by hydroxyl radicals.
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