Transcription factor TaPIL1-mediated iron accumulation in wheat.

Iron (Fe) is an essential micronutrient that helps prevent iron deficiency anemia in humans. To address this problem, iron biofortification represents a promising strategy for developing iron-rich wheat cultivars. In this study, we identified the bHLH transcription factor TaPIL1 as a critical regulator of the wheat response to iron deficiency. Under Fe-deficient conditions, the expression level of TaPIL1 was increased. TaPIL1-5D overexpression (OE) lines accumulated higher iron contents in roots, shoots, and white flour derived from grain than wild type (WT), whereas the tapil1 CRISPR mutants accumulated less iron than WT. Meanwhile, the Fe storage-related genes NATURAL RESISTANCE-ASSOCIATED MACROPHAGE PROTEIN 3 (NRAMP3) and Ferritin1 (Fer1) exhibited higher expression levels in OE lines than in WT. Yeast one-hybrid, EMSA, and LUC activity assays showed that TaPIL1-5D bound to the promoters of TaNRAMP3 and TaFer1 and activated their expression. TaPIL1-5B enhanced the transcriptional activation activity of TaPIL1-5D toward TaNRAMP3 and TaFer1. Moreover, seedlings and grains of wheat varieties carrying the elite haplotype of TaPIL1-5B exhibited higher expression levels of TaNRAMP3 and TaFer1, as well as higher iron contents, than those of wheat varieties carrying the non-elite haplotype. Collectively, our results highlight that the TaPIL1-TaNRAMP3/TaFer1 module functions in iron accumulation in wheat, demonstrating its potential utility for improving iron accumulation in wheat breeding.