Paralogs and mutants show that one DMA synthase functions in iron homeostasis in rice

Rice (Oryza sativa) secretes 2'-deoxymugineic acid (DMA) to acquire insoluble iron (Fe) from the rhizosphere. In rice, DMA is synthesized by DMA synthase 1 (OsDMAS1), a member of the aldo-keto reductase super family. We screened OsDMAS1 paralogs for DMA synthesis. None of these paralogs displayed in vitro DMA synthesis activity, suggesting that rice only harbors one functional DMAS. We further characterized OsDMAS1 mutant plants. We failed to screen homozygous knock-out plants (dmas-1), so we characterized DMAS knock-down plants (dmas-kd1 and dmas-kd2). Under Fe-deficient conditions, dmas-kd1 plants were more chlorotic compared to the wild-type (WT) plants, and the expression of OsNAS3, OsYSL2, OsIRT1, and OsIRO2 was significantly up-regulated in the dmas-kd1 mutant, indicating that metal homeostasis was significantly disturbed. The secretion of DMA in dmas-kd1 was not significantly reduced. The dmas-kd1 plants accumulated less Fe in their roots compared to WT plants when grown with 10 μM FeSO4. The dmas-kd1 plants accumulated more Zn in their roots compared to WT plants under Fe-deficient, Fe-EDTA, and FeSO4 conditions. In both dehusked rice seeds (brown rice) and polished rice, no differences were observed for Fe, Cu, or Mn accumulation, whereas dmas-kd1 seeds significantly accumulated more Zn in brown rice. Our data suggests that rice only harbors one functional gene for DMA synthesis. In addition, the knock-down of OsDMAS1 significantly up-regulates the genes involved in Fe uptake and homeostasis.