Abstract

Iron (Fe) deficiency is a globally widespread condition in which the body lacks sufficient Fe to produce hemoglobin. However, major food crops generally have low grain Fe contents. Consequently, enhancing grain Fe concentrations is important for improving the health of populations that rely on grains as staple foods. Here, we isolated a yellow stripe leaf mutant of foxtail millet ( Setaria italica ), designated yellow stripe-like 1 ( ysl1 ). This mutant exhibited typical Fe deficiency symptoms that were alleviated when grown under Fe-sufficient conditions. Compared with the wild-type, Siysl1 showed lower Fe concentrations in seedling roots, shoots, stems, elongation-stage leaves, panicles, and seeds, but a higher Fe concentration in heading-stage leaves. Using MutMap+, we identified and cloned SiYSL1 and validated its function through CRISPR/Cas9-mediated knockout experiments. SiYSL1 encodes an Fe-phytosiderophore transporter and is highly induced under Fe deficiency conditions. Histochemical staining revealed that SiYSL1 is specifically expressed in vascular bundles of roots and leaves of plants grown under Fe deficiency conditions, and in spikelets, expanding ovaries, basal endosperm, and embryo-surrounding tissues. Thus, SiYSL1 appears to regulate Fe uptake and homeostasis, and plays an essential role in Fe translocation to seeds. The overexpression of SiYSL1 in rice and foxtail millet significantly increased seed Fe contents, suggesting its value in crop breeding. Predicted transcription factor binding sites in the SiYSL1 promoter and a spikelet transcriptome analysis indicated that transcription factors regulate SiYSL1 expression. Our study provides new genetic resources for the Fe bio-enhancement of food crops and insights into the mechanisms responsible for seed Fe accumulation.