Interplay of chelating and reducing root exudates in plant iron acquisition
Final Report Abstract
Under iron deficiency, dicotyledonous plants release coumarin derivatives in their root exudates that are able to mobilize sparingly soluble Fe in the rhizosphere. At the start of this project, only scopoletin was confirmed as a root exudate relevant for Fe mobilization, while other coumarin derivatives and their functional relevance as siderophores were largely unknown. Thus, goals of this project were to identify new coumarin-type siderophores in Arabidopsis, describe their modifications upon release, determine their role in Fe(III)-chelation versus -reduction and describe their efficacy in dependence of soil properties or growth conditions, esp. pH. During the course of this project, which was a cooperation between two German and two Austrian partners in frame of a DACh proposal, the German partners made the following achievements: i) The biosynthesis pathway of Fe-mobilizing coumarins includes two further steps, the hydroxylation of scopoletin to fraxetin by scopoletin-8-hydroxylase, a 2-oxoglutarate-dependent dioxygenase, and the subsequent hydroxylation of fraxetin to sideretin by the cytochrome P450 enzyme CYP82C4. Both enzymes are induced under Fe deficiency and co-localize in epidermal cells. Notably, only s8h but not cyp82C4 mutants display severe Fe deficiency symptoms when grown in soil or agar with low Fe availability. In-vitro assays demonstrated that under sterile conditions scopoletin is ineffective in Fe mobilization, while sideretin mobilizes Fe at slightly acidic pH and only fraxetin is able to mobilize Fe efficiently at high pH, suggesting that different coumarins facilitate Fe acquisition according to the prevailing soil conditions. In-vitro assays showed further that fraxetin and sideretin can reduce Fe(III) with slightly distinct kinetics. Mass spectrometry-based analysis identified new, partly oxidized coumarin species including dimers, hydroxylated dimers, quinones, and demethylated products, while intact coumarin-Fe-complexes could be identified unequivocally but not yet quantified in root exudates. Model experiments proposed a pathway, by which scopoletin can be demethylated in the presence of Fe to yield esculetin, another catechol-type coumarin. Growth assays with single and double mutants defective in individual steps of coumarin biosynthesis and Fe(III) reduction provided direct genetic support for the role of coumarins in Fe(III) reduction with fraxetin being most effective at neutral to alkaline pH and sideretin only at slightly acidic pH. Moreover, external medium pH altered the composition of coumarins in root exudates, partly through transcriptional regulation of coumarin biosynthesis genes and in dependency of a functional Fe(III) reductase. Taken together, the results of this project show that the biosynthesis and release of coumarins underlie high plasticity, by which plants adapt to the external conditions, under which Fe chelation and reduction have to take place.
Publications
- (2018) Biosynthesis of redox-active metabolites in response to iron deficiency in plants. Nature Chemical Biology 14, 442-450
Rajniak J., Giehl R.F.H., Chang E., Murgia I., von Wirén N., Sattely E.S.
(See online at https://doi.org/10.1038/s41589-018-0019-2) - (2020) Importance of oxidation products in coumarin-mediated Fe(hydr)oxide mineral dissolution. BioMetals 33, 305-321
Baune M., Kang K., Schenkeveld W.D.C., Kraemer S.M., Hayen H., Weber G.
(See online at https://doi.org/10.1007/s10534-020-00248-y) - (2021) Root exudation of coumarins from soil-grown Arabidopsis thaliana in response to iron deficiency. Rhizosphere 17, 100296
Rosenkranz T., Oburger E., Baune M., Weber G., Puschenreiter M.
(See online at https://doi.org/10.1016/j.rhisph.2020.100296)
