Biology subjects

Cotelle, V.

Publications and source records attributed to Cotelle, V..

3 recordsLinked to original sources

bioRxiv2026

TurboID-based proteomic profiling reveals proxitome of the IRT1 metal transporter and new insight into metal uptake regulation in plants

IRT1 is the major root iron transporter responsible for broad spectrum metal absorption in Arabidopsis root epidermal cells. Non-iron metal substrates of IRT1 were recently shown to regulate IRT1 cell surface levels by endocytosis and vacuolar degradation. TurboID-based proximity labeling was recently developed to detect protein:protein interactions and thus shed light on the intricate regulation of proteins in living cells. Although TurboID-based proximity labeling technology has been successfully established in mammals, its application in plant systems remains limited and inexistent for highly hydrophobic multispann transmembrane proteins. Here, we established TurboID for proximity labeling of IRT1 and identified 494 IRT1-specific proximal proteins, including the previously reported FYVE1 IRT1 interacting protein. To showcase the biological relevance of identified IRT1 proximal proteins, we characterized further the NHX5 Na+(K+)/H+ antiporter and the RGLG2 E3 ubiquitin ligase. We validated both IRT1 proximal proteins as IRT1 partners using several orthogonal assays. We also highlight the contribution of NHX-type antiporters and RGLG-type E3 ligases in plants responses to non-iron metal nutrition and IRT1 endocytosis. Overall, our work showcases the power of TurboID to identify new interacting proteins for plant transporters, expanding the application of this technology to proteins notoriously difficult to work with.

plant biology↗

bioRxiv2024

Metal sensing properties of the disordered loop from the Arabidopsis metal transceptor IRT1

The plant IRT1 iron transporter is a plasma membrane protein that takes up iron in root upon iron-limited conditions. Besides its primary metal substrate iron, IRT1 also transports other divalent metals that overaccumulate in plants when soil iron is low and IRT1 is highly expressed. We previously reported that the intracellular regulatory loop between transmembrane helices TM4 and TM5, comprising IRT1 residues from 144 to 185, is involved in the post-translational regulation of IRT1 by its non-iron metal substrates. Upon excess of zinc, IRT1 (144-185) undergoes phosphorylation by the CIPK23 kinase followed by its ubiquitination by IDF1 to target IRT1 for vacuolar degradation. This zinc-dependent downregulation of IRT1 requires the presence of four histidine (H) residues in IRT1 loop, that directly bind zinc. However, how selective metal binding is achieved and how this allows downstream regulation to take place is largely unknown. Here, we characterized the metal binding properties and structure of IRT1 loop to better understand the molecular basis of non-iron metal sensing and signaling. Using a combination of circular dichroism and NMR, we demonstrate that zinc and manganese bind to IRT1 loop with nanomolar range affinity, and that metal binding does not trigger structuration of the loop. We prove that zinc and manganese binding is mediated by the four H residues and identify aspartic acid (D) residue D173 as helping in metal coordination and participating to metal sensing and metal-dependent degradation of IRT1 in plants. Altogether, our data provide further evidence of how the regulatory loop of IRT1 senses high cytosolic divalent metal concentrations to regulate metal uptake in plants.

plant biology↗

bioRxiv2023

Interdependence of a kinase and its cognate substrate plasma membrane nanoscale dynamics underlies Arabidopsis response to viral infection

Plant viruses represent a risk to agricultural production and as only a few treatments exist, it is urgent to identify resistance mechanisms and factors. In plant immunity, plasma membrane (PM)-localized proteins play an essential role in sensing the extracellular threat presented by bacteria, fungi or herbivores. Viruses are intracellular pathogens and as such the role of the plant PM in detection and resistance against viruses is often overlooked. We investigated the role of the partially PM-bound Calcium-dependent protein kinase 3 (CPK3) in viral infection and we discovered that it displayed a specific ability to hamper viral propagation over CPK isoforms that are involved in immune response to extracellular pathogens. More and more evidence support that the lateral organization of PM proteins and lipids underlies signal transduction in plants. We showed here that CPK3 diffusion in the PM is reduced upon activation as well as upon viral infection and that such immobilization depended on its substrate, Remorin (REM1.2), a scaffold protein. Furthermore, we discovered that the viral infection induced a CPK3-dependent increase of REM1.2 PM diffusion. Such interdependence was also observable regarding viral propagation. This study unveils a complex relationship between a kinase and its substrate that contrasts with the commonly described co-stabilisation upon activation while it proposes a PM-based mechanism involved in decreased sensitivity to viral infection in plants.

plant biology↗