Lab manager/Engineer CNRS
Associate Professor UPS
Reyes RODENAS CASTILLO
Alvaro Montiel Jorda, PhD
Claire Simon Vezo, Master
Amanda Martin Barranco, PhD
Guillaume Dubeaux, PhD
Sara Martins PhD
Natali Romero Barrios, postdoc
Alex Johnson, postdoc
Enric Zelazny, postdoc, CRCN CNRS
Anne Cayrel, Lab manager CNRS
Marie Barberon, PhD
Alicia Sivitz, postdoc
Robert Blanvillain, postdoc
,Plant cells use a network of signaling pathways to translate information from endogenous programs and environmental conditions into changes in growth or identity. We use the model plant Arabidopsis thaliana to dissect the regulatory mechanisms allowing plants to fine tune signaling pathways underlying endogenous (hormones) and exogenous cues (soil nutrient concentrations, light, temperature) to create robust responses integrating several signals and regulatory levels.
The Vert lab focuses on the role of ubiquitin using plants as models. The post-translational modification ubiquitination involves the covalent linkage of ubiquitin moieties on a lysine residue of a target protein. Ubiquitination exists under different forms, ranging from the addition of one ubiquitin (monoubiquitination) to the formation of polyubiquitin chains. Several types of polyubiquitin chains are found, depending on the residue from ubiquitin engaged in the chain formation. The topologies displayed by the different polyubiquitin chains are associated with various biological outputs, including degradation by the proteasome, ER-associated degradation, endocytosis, DNA repair, transcription activation, etc. As such, ubiquitination is involved in a wide array of cellular processes and signaling pathways. Our group is mostly focused on proteasome- independent functions of ubiquitin, particularly in the context of monoubiquitin and polyubiquitin chains involving the lysine residue K63 from ubiquitin. K63 polyubiquitin-linked chains represent the second most abundant form of ubiquitin-based post-translational modification.
The role of these sub-types of ubiquitination events on the regulation of protein activity (levels, dynamics, activity per se) and signaling pathways are studied using Arabidopsis thaliana as a multicellular organism. The Lab is more specifically interested in :
1) understanding how plants uses the post-translational modification ubiquitination to control the localization and abundance of intrinsic plasma membrane proteins in response to developmental and environmental cues, and
2) unraveling the networks of ubiquitination and discovering new proteasome-independent roles for ubiquitination using plants as model. To do so, we use a combination of molecular genetics, high-resolution imaging and proteomics to characterize the machinery driving lysine(K)63-linked polyubiquitin chain formation and its role in plant cells.
For more detail on our work, please check out the following sections, or the Vert lab website.
The levels and the dynamics of many proteins is tightly regulated to rapidly adjust to ever changing environmental conditions. To date, only a few plant plasma membrane proteins were shown to undergo endocytosis, and the molecular determinants of their dynamics are largely unknown. Our group is interested in studying the dynamics of the Arabidopsis IRT1 transporter as a model for endocytosis in plants. We uncovered a role for monoubiquitination of IRT1 in the specific control of IRT1 levels at the plasma membrane by endocytosis (Barberon et al., 2011). We demonstrated that the ubiquitination of IRT1 is regulated by certain environmental signals. The multi-level integration of signals based on transcriptional and post-translational regulations thus allow a tight control of iron uptake in time and space. We are now investigating the existence, the cellular and physiological roles and the mechanisms of plasma membrane protein ubiquitination to other cell-surface receptors and transporters (Martins et al., 2015). Notably, we are now searching for the machinery driving ubiquitination and ubiquitin- dependent endocytosis for plasma membrane proteins using IRT1 and receptor kinases such as the steroid hormone receptor BRI1 and developing high- and super-resolution microscopy to better understand their dynamics.
Molecular mechanisms of K63 polyUb-dependent endocytosis
We previously demonstrated that the endocytosis and vacuolar degradation of IRT1 and BRI1 was controlled by their monoubiquitination and K63 polyubiquitination, respectively (Barberon et al., 2011 ; Martins et al., 2015). Using a combination of confocal microscopy and high resolution TIRF imaging, we unravelled the dual role of ubiquitin in endocytosis. Cargo ubiquitination playes a partial role in internalization from the cell surface but is absolutely essential for proper sorting and targeting to the vacuole. Besides, we showed that IRT1 and BRI1 ubiquitination was a critical process controlling the amount of both proteins in the cell and thereby controlling plant metal uptake and responses to steroid hormones (Barberon et al., 2011 ; Barberon et al., 2014 ; Dubeaux et al., 2018 ; Martins et al., 2015 ; Martins et al., 2017).
Influence of environmental conditions on K63 polyubiquitin-dependent endocytosis
We are now investigating how environmental conditions are impinging on IRT1 and BRI1 protein dynamics and levels through differential ubiquitination. We already demonstrated that secondary metal substrates of IRT1 regulate IRT1 localization and levels through ubiquitin-mediated endocytosis (Barberon et al., 2014 ; Dubeaux et al., 2018). We have also identified environmental conditions that modulate the amount and localization of BRI1 via its ubiquitination, thus impacting on steroid-dependent growth (Martins et al., 2017). Our objective is now to identify the factors underlying such regulation and evaluate their contribution in plant growth and development.
High resolution imaging of endocytosis and role of K63 polybiquitination
We have implemented TIRF microscopy imaging of Arabidopsis roots and hypocotyls to monitor single endocytosis events at the cell surface (Martins et al., 2015 ; Johnson et al., 2017). We have determined the lifetime at the plasma membrane of cargos and endocytosis-related proteins and are now evaluating how ubiquitination of cargo proteins influence the recruitment of the endocytic machinery and vesicle formation. An example of TIRF microscopy image of the endocytic adaptor AP2 fused to GFP is shown below. Individual endocytic events are oberseved and semi-automatically detected to reveal its cell surface residence time.
The second objective of the lab is to identify and characterize the machinery driving the formation of K63 polyubiquitin chains in plants. The completion of the Arabidopsis genome sequencing project revealed the dramatic expansion of the ubiquitination machinery in plants, with over 1,400 genes coding factors involved in ubiquitination processes (>5% of the genome). We therefore initiated several complementary approaches using genetics, proteomics and genomics to tackle the complexity of ubiquitination processes in higher eukaryotes and to highlight new biological functions of K63 polyubiquitination.
Using a sensor named Vx3K0 that specifically recognizes K63 polyubiquitin chains, we isolated for the first time proteins modified with K63 chains and identified these proteins by mass spectrometry (Johnson and Vert, 2016 ; Romero-Barrios et al., 2020). This provides a new resolution in our understanding of ubiquitination processes and allows us to focus on the roles specifically associated with K63 polyubiquitination. Besides numerous plasma membrane proteins recognized by Vx3K0, we also identified several interesting cytosolic and nuclear proteins decorated with K63-linked polyubiquitin chains and are currently addressing their functional importance. In parallel, we isolated E3 ligases involved in substrate selection for K63 polyubiquitination using large-scale yeast two hybrid screening using the K63 polyubiquitin-dedicated E2 enzymes UBC35 and UBC36 as well as their cognate E2 variants as bait. We notably identified several E3/substrate couples, offering new insight into the mechanisms of K63 polyubiquitin chain formation and its biological roles (Johnson and Vert, 2016 ; Romero-Barrios et al., 2020).
The SWELL lines are a collection of markers expressed in specific root tissues and that allows functional genomic in Arabidopsis thaliana, including tissue specific gene induction, nuclei purification by the INTACT method and multicolor imaging (Marques-Bueno et al., 2016).
Platre, M., Satbhai, S., Brent, L., Gleason, M., Cao, M., Grison, M., Glavier, M., Zhang, L., Gaillochet, C., Göschl, C., Giovannetti, M., Enugutti, B., Neveu, J., von Reth, M., Alcázar, R., Parker, J., Vert, G., Bayer, E., and Busch, W. (2022). The receptor kinase SRF3 coordinates iron-level and flagellin dependent defense and growth responses in plants. Nature Communications 13(1):4445. Pubmed link
Martin-Barranco, A., Vert, G., Thomine, S. and Zelazny, E. (2021). A quick journey into the diversity of iron uptake strategies in photosynthetic organisms. Plant Signaling and Behavior 16(11):1975088. Pubmed link
Rodenas, R. and Vert, G. (2021). Regulation Of Root Nutrient Transporters By CIPK23: « One Kinase To Rule Them All ». Plant Cell Physiology 62(4):553-563. Pubmed link
Ivanov, R. and Vert, G (2021). Endocytosis in plants: peculiarities and roles in the regulated trafficking of plant metal transporters. Biology of the Cell 113(1):1-13. Pubmed link
Spielmann, J. and Vert, G (2021). The many facets of protein ubiquitination and degradation in plant root iron deficiency responses. Journal of Experimental Botany 72(6):2071-2082. Pubmed link
Martin Barranco, A., Spielmann, J., Dubeaux, G., Vert, G. and Zelazny, E. Dynamic control of the Arabidopsis high affinity iron uptake complex in root epidermal cells. Plant Physiology doi: 10.1104/pp.20.00234. Pubmed link
Liu, D., Kumarn R., Claus, L.A.N., Johnson, A., Vanhotte, I., Wang, P., Bender, K., Yperman, K., Martins, S., Zhao, X., Vert, G., Van Damme, D., Friml, J. and Russinova, E. (2020). Endocytosis of BRASSINOSTEROID INSENSITIVE1 is partly driven by a canonical tyrosine-based motif. Plant Cell 32, 3598-3612. Pubmed link
Johnson, A., Gnyliukh, N., Kaufmann, W., Narasimhan, M., Vert, G., Bednarek, S., and Friml, J. (2020). Experimental toolbox for quantitative evaluation of clathrin mediated endocytosis in the plant model Arabidopsis. Journal of Cell Science 133:jcs248062. Pubmed link
Vert, G. (2020). Plant Cell Signaling: SUMO is Under the Influence of Steroids and Salt. Current Biology 30, 342-344. Pubmed link
Romero-Barrios, N., Monachello, D., Dolde, U., Cayrel, A., Wong, A., San Clemente, H., Johnson, A., Lurin, C. and Vert, G. (2020). Advanced cataloging of K63 polyubiquitin networks by genomic, interactome and sensor-based proteomic analyses. Plant Cell 32, 123-138. Pubmed link
Cointry, V. and Vert, G. (2019). The bifunctional transporter-receptor IRT1 at the heart of metal sensing and signaling. New Phytologist 223, 1173-1178. Pubmed link
Mishev, K., Lu, Q., Denoo, B., Peurois, F., Dejonghe, W., Hullaert, J., Sharma, I., Nerinckx, W., De Rycke, R., Goodman, K., Kalinowska, K., Storme, V., Boeren, S., Son Long Nguyen, L., Drozdzecki, A., De Munck, S., Martins, S., Vert, G., Audenaert, D., Otegui, M., Isono, E., Madder, A., De Vries, S., Savvides, S., Cherfils, J., Winne, J., and Russinova, E. (2018). Nonselective chemical inhibition of Sec7 domain-containing ARF GEFs in Arabidopsis. Plant Cell 30, 2573-2593. Pubmed link
Dubeaux, G., Neveu, J., Zelazny, E. and Vert, G. (2018). Metal sensing by the IRT1 transporter/receptor orchestrates its own degradation and plant metal nutrition. Molecular Cell 69, 953-964. Pubmed link
Martins, S., Montiel-Jorda, A., Cayrel, A., Huguet, S., Paysant-Le Roux, C., Ljung, K., and Vert, G. (2017). Brassinosteroid signaling-dependent root responses to prolonged elevated ambient temperature. Nature Communications 8, 309. Pubmed link
Dubeaux, G. and Vert, G. (2017). Zooming into plant ubiquitin-mediated endocytosis. Current Opinion in Plant Biology 40, 56- 62. Pubmed link
Johnson, A. and Vert, G. (2017). Single event resolution of plasma membrane protein endocytosis by TIRF microscopy. Frontiers in Plant Science 8, 612. Pubmed link
Martins, S. Vert, G. and Jaillais, Y. (2016). Probing activation/deactivation of the BRASSINOSTEROID INSENSITIVE1 receptor kinase by immunoprecipitation. Methods in Molecular Biology 1564, 169-180. Pubmed link
Jaillais, Y. and Vert, G. (2016). Brassinosteroid signaling and BRI1 dynamics went underground. Current Opinion in Plant Biology 33, 92-100. Pubmed link
Johnson, A. and Vert, G. (2016). Unravelling K63 polyubiquitination networks by sensor-based proteomics. Plant Physiology 171, 1808-1820. Pubmed link
Wild, M., Davière, J.M., Regnault, T., Sakvarelidze-Achard, L., Carrera, E., Lopez Diaz, I., Cayrel, A., Dubeaux, G., Vert, G., Achard, P. (2016).Tissue-specific regulation of gibberellin signaling fine-tunes Arabidopsis iron deficiency responses. Developmental Cell 37,190-200. Pubmed link
Marquès-Bueno, M. Morao, A.K., Cayrel, A., Platre, M., Barberon, M., Cailleux, E., Colot, V., Jaillais, Y., Roudier, F., and Vert, G. (2016) A versatile multisite Gateway-compatible promoter and transgenic line collection for cell type-specific functional genomics in Arabidopsis. Plant Journal 85, 320-333. Pubmed link
Zelazny, E. and Vert, G. (2015). Regulation of iron uptake by IRT1: Endocytosis pulls the trigger. Molecular Plant 8, 977-979. Pubmed link
Martins, S., Dohmann, E., Cayrel, A., Johnson, A., Fischer, W., Pojer, F., Satiat-Jeunemaitre, B., Jaillais, Y., Chory, J., Geldner, N. and Vert, G. (2015). Internalization and vacuolar targeting of the brassinosteroid hormone receptor BRI1 are regulated by ubiquitination. Nature Communications 6, 6151. Pubmed link
Dubeaux, G., Zelazny, E. and Vert, G. (2015). Getting to the root of plant iron uptake and cell-cell transport : Polarity matters !Communicative and Integrative Biology 8, e1038441. Pubmed link
Zelazny, E. and Vert, G. (2014). Plant nutrition : root transporters on the move. Plant Physiology 166, 500-508. Pubmed link
Luschnig, C., and Vert, G. (2014). The dynamics of plant membrane proteins: PINs and beyond. Development 141, 2924-2938. Pubmed link
Barberon, M., Dubeaux, G., Kolb, C., Isono, E., Zelazny, E., and Vert, G. (2014). Polarization of IRON-REGULATED TRANSPORTER 1 (IRT1) at the plant-soil interface plays crucial role in metal homeostasis. PNAS 111, 8293-8298. Pubmed link
Thomine, S., and Vert, G. (2013). Iron transport in plants : better be safe than sorry! Current Opinion in Plant Biology 16, 322-327. Pubmed link
Sivitz, A., Hermand, V., Curie, C. and Vert, G. (2012). Arabidopsis bHLH100 and bHLH101 control iron homeostasis via a FIT- independent pathway. PLoS One 7, e44843-e44854. Pubmed link
Jaillais, Y. and Vert, G. (2012). Brassinosteroids, gibberellins and light-mediated signalling are the three-way controls of plant sprouting. Nature Cell Biology 14, 788-790. Pubmed link
Vert, G., and chory, J. (2011). Crosstalk in Cellular Signaling : Background noise or the Real Thing? Developmental Cell 6, 985- 991. Pubmed link
Zelazny, E., Barberon, M., Curie, C., and Vert, G. (2011). Ubiquitination of transporters at the forefront of plant nutrition. Plant Signaling and Behavior 10, 1597-1599. Pubmed link
Roschzttardtz, H., Seguela, M., Briat, J.F., Vert, G., and Curie, C. (2011). The citrate effluxer FRD3 facilitates mineral nutrition during post-germinative seedling and pollen development. Plant Cell 7, 2725-2737. Pubmed link
Barberon, M., Zelazny, E., Robert, S., Conejero, G., Curie, C., Friml, J., and Vert, G. (2011). Author Summary : Monoubiquitin- dependent endocytosis of the root transporter IRT1 controls iron uptake in plants. PNAS PLUS 108, 12985-12986. PNAS link
Barberon, M., Zelazny, E., Robert, S., Conejero, G., Curie, C., Friml, J., and Vert, G. (2011). Monoubiquitin-dependent endocytosis of the root transporter IRT1 controls iron uptake in plants. PNAS PLUS 108, E450-E458. Pubmed link
Vert, G. and Chory, J. (2009). A Toggle Switch in Plant Nitrate Uptake. Cell 138, 1064-1066. Pubmed link
De Rybel, B., Audenaert, D., Vert, G., Rozhon, W., Mayerhofer, J., Peelman, F., Coutuer, S., Denayer, T., Jansen, L., VanHoutte, I., Beemster, G., Vleminckx, K., Jonak, C., Chory, J., Inzé, D., Russinova, E and Beeckman, T. (2009). Chemical inhibition of a subset of Arabidopsis thaliana GSK3-like kinases activates brassinosteroid signaling. Chemistry and Biology 16, 594-604. Pubmed link
Vert, G. (2008). Plant signaling: brassinosteroids, immunity and effectors are BAK ! Current Biology 18, 963-965. Pubmed link
Vert, G., Walcher, C., Chory, J., and Nemhauser, J. (2008). Integration of auxin and brassinosteroid pathways by Auxin Response Factor 2. PNAS 105, 9829-9834. Pubmed link
Briat, J.F., Arnaud, N., Cellier, F., Curie, C., Gaymard, F., Ravet, K., Séguéla, M., and Vert, G. (2007). Iron uptake and storage in plants. American Journal of Hematology 82, 506-507.
Vert, G. and Chory, J. (2006). Downstream nuclear events in brassinosteroid signaling. Nature 441, 96-100. Pubmed link
Schaaf, G., Schikora, A., Häberle, J., Vert, G., Ludewig, U., Briat, J.-F., Curie, C., and Von Wirén, N. (2005). A putative function of the ArabidopsisFe-phytosiderophore transporter homolog AtYSL2 in Fe and Zn homeostasis. Plant Cell Physiology 46, 762-774. Pubmed link
(Mora-Garcia, S., Vert, G.) co-first authors, Yin, Y., Cano-Delgado, A., Cheong, H., and Chory, J. (2004). Nuclear protein phosphatases with Kelch-repeat domains modulate the response to brassinosteroids in Arabidopsis. Genes & Dev. 18,, 448-460.
Pubmed linkVansuyt, G., Vert, G., and Larcher, M. (2005). Les régulateurs de croissance en agriculture. In : Regnault-Roger, C. Enjeux phytosanitaires pour l’agriculture et l’environnement du XXI° siècle. Tec et Doc Lavoisier, Paris.
Briat J.-F. and Vert, G. (2004). Acquisition et gestion du fer par les plantes. Cahiers/Agricultures 13, 183-201.
Vert, G., Briat, J.-F., and Curie, C. (2003). Dual regulation of the Arabidopsis high affinity root iron uptake system by local and long-distance signals. Plant Physiology 132, 796-804. Pubmed link
Vert, G., Grotz, N., Dedaledechamp, F., Gaymard, F., Guerinot, M.L., Briat, J.-F., and Curie, C. (2002). IRT1: an Arabidopsis iron transporter essential for iron uptake from the soil and plant growth. Plant Cell 14, 1223-1233. Pubmed link
Vert, G., Briat, J.-F., and Curie, C. (2001). Arabidopsis IRT2 gene encodes a root periphery iron transporter. Plant Journal 26, 181-189. Pubmed link
C. DELESALLE, University Paul Sabatier Toulouse 3
F. DELIGNE, University Paul Sabatier Toulouse 3
V. COINTRY (2021), University Paris-Saclay
A. MARTIN-BARRANCO (2020), University Paris-Saclay
A. MONTIEL-JORDA (2019), University Paris-Saclay
G. DUBEAUX (2016), University Paris-Saclay
S. MARTINS (2016), University Paris-Saclay
M. BARBERON (2010). University Montpellier 2
M. SEGUELA (2007). University Montpellier 2
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