Iron (Fe) is an essential cofactor for many cellular redox reactions and is required for life. Plants tightly regulate Fe uptake to maintain a balance between the demand for Fe in photosynthetic tissues and the risk of over-accumulation that could lead to cellular damage. Although the mechanisms of root Fe uptake and the transcriptional networks that control root-level regulation of Fe uptake have been well-studied, the mechanisms by which shoots signal Fe status to the roots still remain unresolved. In plants, none of the metal sensing or signaling systems is completely understood to date. Earlier research in several labs, including our own, has suggested the existence of long-distance shoot signals that strongly control Fe uptake by roots. While conducting seedling grafting experiments, we realized that excised roots might provide a simple bioassay for a mobile signal moving in the phloem. We developed an assay that tested whether phloem exudates collected from Fe-sufficient (+Fe) and Fe-deficient (-Fe) plants are able to induce Fe-deficiency-regulated gene expression in wild-type (WT) roots. We found that a substance present only in phloem exudate from –Fe plants is a positive signal that activates Fe-deficiency-regulated gene expression in the roots. Characterization of the signal indicated that the signal is around 10 kDa and is a small RNA (sRNA). sRNA sequencing data suggest that an Fe-regulated, long non-coding RNA might be the precursor of a signal that positively regulates Fe uptake by the roots. Improved understanding of the mechanism of long-distance Fe signaling will allow improved strategies for the engineering of staple crops to accumulate additional bioavailable Fe in edible parts, thus improving the Fe nutrition of the billions of people worldwide whose inadequate diet causes Fe-deficiency anemia. Keywords: Iron uptake, iron deficiency, phloem exudate, signal, sRNA, long non-coding RNA.