Membrane microdomain HIR proteins: control of iron uptake machinery and new functions in Arabidopsis – NUTRIR

- High resolution microscopy
- Biochemistry
- Interactome
- Protein-protein interaction test
- CRISPR/Cas9 technology
- Phenotypical analysis

Iron is essential for plant growth and development, by playing fundamental roles in respiration and photosynthesis. Iron absorption in root epidermal cells is achieved by the IRT1 iron transporter that also allows the entry of non-iron metals such as Zn, Mn and Co. As recently demonstrated, IRT1 endocytosis is controlled by non-iron metals through IRT1 ubiquitination processes. We established an interactome of IRT1 and discovered that the AHA2 proton pump and the FRO2 reductase, both of which work in concert with IRT1 in the strategy of acidification-reduction-transport of iron, directly interact with IRT1. The study of this iron-acquisition complex showed that the ubiquitination of FRO2 and AHA2 is independent of the non-iron metal status and that these proteins are not massively endocytosed in response to an excess of these metals, contrary to IRT1. Indeed, our results suggest that the phosphorylation of IRT1 in response to an excess of non-iron metals induces the dissociation of the IRT1/FRO2/AHA2 complex. We propose that an iron-acquisition complex exists at the surface of root epidermal cells to optimize iron acquisition in plants (Martin-Barranco et al 2020). In addition, this “iron acquisition platform” may be regulated through a recruitment in specific membrane nanodomains by interacting with HIR2. This hypothesis is currently studied.

.

Martin-Barranco A, Spielmann J, Dubeaux G, Vert G, Zelazny E (2020) Dynamic Control of the High-Affinity Iron Uptake Complex in Root Epidermal Cells. Plant Physiology 184: 1236-1250. Référence HAL : hal-02931857v1.

Membrane microdomains are specifically enriched in sterols, sphingolipids and specific subsets of proteins such as the stomatin/prohibitin/flotillin/HflK/C (SPFH) domain-containing proteins that are found in most evolutionary lineages. Interestingly, plants possess a specific group of SPFH domain-containing proteins named Hypersensitive Induced Reaction proteins (HIRs) composed of four members (HIR1 to HIR4). Although in animals SPFH proteins are involved in essential cellular processes such as the regulation of membrane protein activity or endocytic mechanisms, the function of HIRs remains largely unknown in plants. The first aim of the NUTRIR project is: (i) to characterize the small Arabidopsis HIR protein family, at the cellular level in terms of localization, dynamics, and microdomain identity to shed light on their role in the cell using cutting-edge microscopy techniques, (ii) to discover new roles of these proteins using HIR2 isoform as a model. For this purpose we will identify HIR2-interacting proteins and investigate the importance of these interactions at the cellular and physiological levels. Recently, I discovered that the Arabidopsis root iron transporter Iron Regulated Transporter1 (IRT1), which is one of the model proteins studied in our team, physically interacts with HIR2. Moreover we showed that IRT1 interacts with the proton pump H+-ATPase2 (AHA2) and the reductase Ferric Reduction Oxydase2 (FRO2) that are also essential for iron acquisition in Arabidopsis roots. Interestingly, we demonstrated by using reverse genetic approaches that HIR2 is involved in the maintenance of metal homeostasis in Arabidopsis. The second aim of the NUTRIR project is to specifically study the role of HIR2 in the regulation of the iron acquisition machinery focusing on its role in the intracellular dynamics of IRT1 and the impact on metal homeostasis.