The regulation of adaptive responses to phosphorus (P) deficiency by the microRNA399 (miR399)/PHOSPHATE2 (PHO2) pathway has been well studied in Arabidopsis thaliana but not in maize (Zea mays). Here, we show that miR399 transcripts are strongly induced in maize by phosphate (Pi) deficiency. Transgenic maize plants that over-expressed MIR399b accumulated excessive amounts of P in their shoots and displayed typical Pi-toxicity phenotypes. We re-annotated ZmPHO2 with an additional 1165 bp of the 59-untranslated region. miR399-guided post-transcriptional repression of ZmPHO2 was mainly observed in the P-efficient lines. We identified Pi-deficiency-induced long non-coding RNA 1 (PILNCR1) from our strand-specific RNA libraries. Transient expression assays in Nicotiana benthamiana and maize leaf protoplasts demonstrated that PILNCR1 inhibits ZmmiR399-guided cleavage of ZmPHO2. The abundance of PILNCR1 was significantly higher in P-inefficient lines than in P-efficient lines, which is consistent with the abundance of ZmmiR399 transcripts. These results indicate that the interaction between PILNCR1 and miR399 is important for tolerance to low Pi in maize.

Iron (Fe) deficiency remains widespread among people in developing countries. To help solve this problem, breeders have been attempting to develop maize cultivars with high yields and high Fe concentrations in the kernels. We conducted a genome-wide association study and identified a gene, ZmNAC78 ( NAM/ATAF/CUC DOMAIN TRANSCRIPTION FACTOR 78 ), that regulates Fe concentrations in maize kernels. We cultivated maize varieties with both high yield and high Fe concentrations in their kernels by using a molecular marker developed from a 42–base pair insertion or deletion (indel) in the promoter of ZmNAC78 . ZmNAC78 expression is enriched in the basal endosperm transfer layer of kernels, and the ZmNAC78 protein directly regulates messenger RNA abundance of Fe transporters. Our results thus provide an approach to develop maize varieties with Fe-enriched kernels.