A Model to Incorporate the bHLH Transcription Factor OsIRO3 within the Rice Iron Homeostasis Regulatory Network.

Iron (Fe) homeostasis in plants is governed by a complex network of regulatory elements and transcription factors (TFs), as both Fe toxicity and deficiency negatively impact plant growth and physiology. The Fe homeostasis network is well characterized in and remains poorly understood in monocotyledon species such as rice ( L.). Recent investigation of the rice Fe homeostasis network revealed OsIRO3, a basic Helix-Loop-Helix (bHLH) TF as a putative negative regulator of genes involved in Fe uptake, transport, and storage. We employed CRISPR-Cas9 gene editing to target the coding sequence and generate two independent T-DNA-free, loss-of-function mutants in rice cv. Nipponbare. The mutant plants had similar phenotype under nutrient-sufficient conditions and had stunted growth under Fe-deficient conditions, relative to a T-DNA free, wild-type control (WT). Under Fe deficiency, mutant shoots had reduced expression of Fe chelator biosynthetic genes ( and ) and upregulated expression of an Fe transporter gene (), relative to WT shoots. We place our results in the context of the existing literature and generate a model describing the role of OsIRO3 in rice Fe homeostasis and reinforce the essential function of OsIRO3 in the rice Fe deficiency response.