Double knockout of rice Vacuolar Iron Transporters OsVIT1 and OsVIT2 reveals a trade-off between iron biofortification and iron excess tolerance

Rice (Oryza sativa L.) is a staple food for half of the world’s population, but lacks essential nutrients such as iron (Fe), contributing to widespread human Fe deficiency. The vacuolar Fe transporters OsVIT1 and OsVIT2 limit Fe translocation to grains by sequestering Fe into vacuoles of vegetative tissues, and single mutants of either gene show elevated grain Fe levels. Rice is cultivated in flooded soils, where soluble Fe²⁺ accumulates to toxic levels, raising the question of whether OsVIT1 and OsVIT2 also contribute to Fe detoxification. To investigate this, we generated osvit1osvit2 double mutants and assessed their impact on grain Fe accumulation and Fe excess tolerance. Double mutants had increased Fe concentrations in brown rice, particularly in the scutellum and plumule, indicating a role for OsVITs in determining spatial Fe distribution within the embryo. Root uptake contributed to elevated Fe in grains, whereas Fe translocation from flag leaves was less important. Notably, osvit1osvit2 plants were hypersensitive to excess Fe, revealing a trade-off between Fe biofortification and tolerance to Fe excess. These findings highlight the need to consider Fe excess tolerance in biofortification strategies using VIT genes, and establish OsVIT1 and OsVIT2 as key components for Fe detoxification.