Dynamic Control of the High-Affinity Iron Uptake Complex in Root Epidermal Cells

Abstract

In plants, iron uptake from the soil is tightly regulated to ensure optimal growth and development. Iron absorption in Arabidopsis root epidermal cells requires the IRT1 transporter that also allows the entry of certain non-iron metals, such as Zn, Mn, and Co.Recent work demonstrated that IRT1 endocytosis and degradation are controlled by IRT1 non-iron metal substrates in an ubiquitin-dependent manner. To better understand how metal uptake is regulated, we identified IRT1-interacting proteins in Arabidopsis roots by mass spectrometry and established an interactome of IRT1. Interestingly, the AHA2 proton pump and the FRO2 reductase, both of which work in concert with IRT1 in the acidificationreduction-transport strategy of iron uptake, were part of this interactome. We confirmed that IRT1, FRO2, and AHA2 associate through co-immunopurification and split-ubiquitin analyses, and uncovered that they form tripartite direct interactions. We characterized the dynamics of the iron uptake complex and showed that FRO2 and AHA2 ubiquitination is independent of the non-iron metal substrates transported by IRT1. In addition, FRO2 and AHA2 are not largely endocytosed in response to non-iron metal excess, unlike IRT1. Indeed, we provide evidence that the phosphorylation of IRT1 in response to high levels of non-iron metals likely triggers dissociation of the complex. Overall, we propose that a dedicated ironacquisition protein complex exists at the cell surface of Arabidopsis root epidermal cells to optimize iron uptake.

Root-to-shoot iron partitioning in Arabidopsis requires IRON-REGULATED TRANSPORTER1 (IRT1)

“…Arabidopsis irt1 mutants have severely chlorotic leaves, are strongly impaired in growth and die before reaching the reproductive stage unless supplemented with Fe chelates. The IRT1 protein forms a complex with the FERRIC REDUCTION OXIDASE2 (FRO2) and AUTOINHIBITED PLASMA MEMBRANE H + -ATPase2 (AHA2) for Fe (+III) solubilization and Fe 2+ uptake into epidermal cells in the root differentiation zone (Martín-Barranco et al, 2020). Besides IRT1, the high-affinity Mn transporter NATURAL RESISTANCE-ASSOCIATED MACROPHAGE PROTEIN1 (NRAMP1) is thought to act as a secondary low-affinity Fe 2+ uptake system (Cailliatte et al, 2010; Castaings et al, 2016).…”

“…This leads to the phosphorylation of serine and threonine residues in the same cytosolic loop of IRT1 by CBL-INTERACTING PROTEIN KINASE23 (CIPK23), thus triggering the extension of monoubiquitination into K63-polyubiquitination mediated by IRT1 DEGRADATION FACTOR1 (IDF1), followed by the vacuolar degradation of IRT1 (Dubeaux et al, 2018; Shin et al, 2013). FRO2 and AHA2 can also be ubiquitinated, but – different from IRT1 – their post-translational modification is independent of non-target secondary IRT1 substrate metal cations (Martín-Barranco et al, 2020).…”

IRONMAN Tunes Responses to Iron Deficiency in Concert with Environmental pH

“…Under acidic conditions, Fe is mainly acquired via enzymatic reduction and taken up through FRO2/IRT1. It can be assumed that CYP82C4 activity and sideretin production is closely associated with the AHA2-FRO2-IRT1 module, a supposition that is corroborated by the extremely close correlation of IRT1 and CYP82C4 expression (http://atted.jp), and the identification of CYP82C4 as a potentially interactor of IRT1 (Martín-Barranco et al, 2020). Sideretin exhibit maximal Fe-mobilizing pH capacity at lower pH ranges when compared to fraxetin (Rajanik et al, 2018; Tsai et al, 2018), implying that sideretin secretion is part of the calcifuge branch.…”

The iron will of the research community: advances in iron nutrition and interactions in lockdown times

“…The field of iron nutrition and interactions in plants has made tremendous progress and continues opening new areas of investigation. Nevertheless, there is a need for a better understanding of the biochemical aspects of iron homeostasis, such as protein interactions in iron uptake ( Martín-Barranco et al , 2020 ) and iron signalling, and how iron is trafficked and incorporated in iron-dependent proteins or photosystems. A community resource to catalogue all iron-binding proteins and their function has recently been set up ( Przybyla-Toscano et al , 2021 ).…”