Sebastien LEON

Sebastien LEON
Institut Jacques Monod | IJM · Membrane trafficking, ubiquitin and signaling group

PhD

About

68
Publications
7,044
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1,942
Citations
Citations since 2017
27 Research Items
926 Citations
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2017201820192020202120222023050100150
2017201820192020202120222023050100150
2017201820192020202120222023050100150
Introduction
I am a group leader at the Institut Jacques Monod, one of the largest research institutes in biology in Paris, France. We study how membrane trafficking is regulated by physiological and metabolic challenges. We focus on the role of signaling pathways, ubiquitin and arrestin-like proteins in this process. Recently, we also became interested in the effect of 2-deoxyglucose, a metabolic inhibitor, in protein trafficking and cellular physiology, and associated resistance mechanisms.
Additional affiliations
April 2016 - present
Paris Diderot University
Position
  • Group Leader
September 2012 - April 2016
Institut Jacques Monod
Position
  • Group Leader
October 2008 - September 2012
Institut Jacques Monod
Position
  • Researcher

Publications

Publications (68)
Article
Full-text available
Endocytosis regulates the plasma membrane protein landscape in response to environmental cues. In yeast, the endocytosis of transporters depends on their ubiquitylation by the Nedd4-like ubiquitin ligase Rsp5, but how extracellular signals trigger this ubiquitylation is unknown. Various carbon source transporters are known to be ubiquitylated and e...
Article
Full-text available
Nutrient availability controls the landscape of nutrient transporters present at the plasma membrane, notably by regulating their ubiquitylation and subsequent endocytosis. In yeast, this involves the Nedd4 ubiquitin ligase Rsp5 and arrestin-related trafficking adaptors (ARTs). ARTs are targeted by signaling pathways and warrant that cargo ubiquity...
Article
Full-text available
After endocytosis, membrane proteins can recycle to the cell membrane or be degraded in lysosomes. Cargo ubiquitylation favors their lysosomal targeting and can be regulated by external signals, but the mechanism is ill-defined. Here, we studied the post-endocytic trafficking of Jen1, a yeast monocarboxylate transporter, using microfluidics-assiste...
Article
Full-text available
Anti-cancer strategies that target the glycolytic metabolism of tumors have been proposed. The glucose analog 2-deoxyglucose (2DG) is imported into cells and, after phosphorylation, becomes 2DG-6-phosphate, a toxic by-product that inhibits glycolysis. Using yeast as a model, we performed an unbiased mass spectrometry–based approach to probe the cel...
Preprint
Full-text available
Most malignant cells display increased glucose absorption and metabolism compared to surrounding tissues. This well-described phenomenon results from a metabolic reprogramming occurring during transformation, that provides the building blocks and supports the high energetic cost of proliferation by increasing glycolysis. These features led to the i...
Article
Full-text available
2-deoxyglucose is a glucose analog that impacts many aspects of cellular physiology. After its uptake and its phosphorylation into 2-deoxyglucose-6-phosphate (2DG6P), it interferes with several metabolic pathways including glycolysis and protein N-glycosylation. Despite this systemic effect, resistance can arise through strategies that are only par...
Preprint
Full-text available
2-deoxyglucose is a glucose analog that impacts many aspects of cellular physiology. After its uptake and its phosphorylation into 2-deoxyglucose-6-phosphate (2DG6P), it interferes with several metabolic pathways including glycolysis and protein N-glycosylation. Despite this systemic effect, resistance can arise through strategies that are only par...
Article
Full-text available
The regulation of nutrient uptake into cells is important, as it allows to either increase biomass for cell growth or to preserve homeostasis. A key strategy to adjust cellular nutrient uptake is the reconfiguration of the nutrient transporter repertoire at the plasma membrane by the addition of nutrient transporters through the secretory pathway a...
Article
Most malignant cells display increased glucose absorption and metabolism compared to surrounding tissues. This well-described phenomenon results from a metabolic reprogramming occurring during transformation, that provides the building blocks and supports the high energetic cost of proliferation by increasing glycolysis. These features led to the i...
Article
Full-text available
How cells adjust nutrient transport across their membranes is incompletely understood. Previously, we have shown that S. cerevisiae broadly re-configures the nutrient transporters at the plasma membrane in response to amino acid availability, through endocytosis of sugar- and amino acid transporters (AATs) (Müller et al., 2015). A genome-wide scree...
Article
Full-text available
How cells adjust nutrient transport across their membranes is incompletely understood. Previously, we have shown that S. cerevisiae broadly re-configures the nutrient transporters at the plasma membrane in response to amino acid availability, through endocytosis of sugar- and amino acid transporters (AATs) (Müller et al., 2015). A genome-wide scree...
Article
Ubiquitylation is a reversible post-translational protein modification that regulates a multitude of cellular processes. Detection of ubiquitylated proteins is often challenging, because of their low abundance. Here, we present NUbiCA, a sensitive protein-fragment complementation assay to facilitate the monitoring of ubiquitylation events in cultur...
Article
Full-text available
Polyubiquitin chains linked via lysine (K) 63 play an important role in endocytosis and membrane trafficking. Their primary source is the ubiquitin protein ligase (E3) Rsp5/NEDD4, which acts as a key regulator of membrane protein sorting. The heterodimeric ubiquitin-conjugating enzyme (E2), Ubc13–Mms2, catalyses K63-specific polyubiquitylation in g...
Preprint
Cancer cells display an altered metabolism with an increased glycolysis and glucose uptake. Anti-cancer strategies targeting glycolysis through metabolic inhibitors have been considered. Particularly, the glucose analogue 2-deoxyglucose (2DG) is imported into cells and phosphorylated into 2DG-6-phosphate, a toxic by-product that inhibits glycolysis...
Article
Lipids and proteins are not evenly distributed within the plasma membrane (PM), but instead segregate laterally into many specialized microdomains whose functional relevance is not clear. In this issue, Busto et al (2018) demonstrate that substrate flux through a nutrient transporter drives the lateral relocation of the transporter between specific...
Article
Full-text available
Phytoplankton growth is limited in vast oceanic regions by the low bioavailability of iron. Iron fertilization often results in diatom blooms, yet the physiological underpinnings for how diatoms survive in chronically iron-limited waters and outcompete other phytoplankton when iron becomes available are unresolved. We show that some diatoms can use...
Article
Full-text available
Yeast cells have a remarkable ability to adapt to nutritional changes in their environment. During adaptation, nutrient-signalling pathways drive the selective endocytosis of nutrient transporters present at the cell surface. A current challenge is to understand the mechanistic basis of this regulation. Transporter endocytosis is triggered by their...
Data
Complete lists of Mmi1 protein partners in wt and mot2∆ cells. Shown are lists of proteins co-purified with Mmi1-TAP in the presence or the absence of RNAses in wild type and mot2∆ cells. An untagged strain was used as a negative control. Protein scores (as provided by the Percolator algorithm), percentages of sequence coverage, numbers of peptides...
Data
Analysis of RNA-sequencing data. Shown are genes with a 1.5 fold increase (0.58496250072 on a log2 scale) in expression in mutants relative to the wild type and a p-value<5E-2, the overlap between mot2∆ and rrp6∆ strains, and functional categories of genes upregulated in mot2∆ cells.
Article
Full-text available
In fission yeast, meiosis-specific transcripts are selectively eliminated during vegetative growth by the combined action of the YTH-family RNA-binding protein Mmi1 and the nuclear exosome. Upon nutritional starvation, the master regulator of meiosis Mei2 inactivates Mmi1, thereby allowing expression of the meiotic program. Here, we show that the E...
Chapter
Full-text available
Ubiquitylation is a reversible posttranslational modification that is critical for most, if not all, cellular processes and essential for viability. Ubiquitin conjugates to substrate proteins either as a single moiety (monoubiquitylation) or as polymers composed of ubiquitin molecules linked to each other with various topologies and structures (pol...
Article
Full-text available
α-arrestins play a key role as trafficking adaptors in both yeast and mammals. The yeast Rim8/Art9 α-arrestin mediates the recruitment of ESCRT (endosomal sorting complex required for transport) to the seven-transmembrane protein Rim21 in the ambient pH signaling RIM pathway. ESCRT is thought to function as a signaling platform that enables the pro...
Data
A table listing yeast strains used in this study is provided in Supplementary file 1. DOI: http://dx.doi.org/10.7554/eLife.03307.033
Article
Full-text available
The length of the ubiquitin chain on a substrate dictates various functional outcomes, yet little is known about its regulation in vivo. The yeast arrestin-related protein Rim8/Art9 is monoubiquitinated in vivo by the Rsp5 ubiquitin ligase. This also requires Vps23, a protein that displays an ubiquitin-E2 variant (UEV) domain. Here, we report that...
Article
Full-text available
Regulation of the cellular volume is fundamental for cell survival and function. Deviations from equilibrium trigger dedicated signaling and transcriptional responses that mediate water homeostasis and volume recovery. Cells are densely packed with proteins, and molecular crowding may play an important role in cellular processes. Indeed, increasing...
Article
Full-text available
In metazoans, proteins of the arrestin family are key players of G-protein-coupled receptors (GPCRS) signaling and trafficking. Following stimulation, activated receptors are phosphorylated, thus allowing the binding of arrestins and hence an "arrest" of receptor signaling. Arrestins act by uncoupling receptors from G proteins and contribute to the...
Article
Full-text available
The ubiquitin system is known to be involved in maintaining the integrity of mitochondria, but little is known about the role of deubiquitylating (DUB) enzymes in such functions. Budding yeast cells deleted for UBP13 and its close homolog UBP9 displayed a high incidence of petite colonies and slow respiratory growth at 37°C. Both Ubp9 and Ubp13 int...
Data
Northern analysis of yeast mRNAs. (A) Autoradiographs of washed filters for RNA extracted from yeast and separated in denaturing agarose gels are presented. Yeast strains were cultured with either glucose or galactose as the carbon source (as indicated above the autoradiographs) at two temperatures, 30°C and 37°C (as indicated above the autoradiogr...
Data
The deletion of UBP9 and UBP13 and the single deletion of DUF1 impair the synthesis of the mitochondrial ATP synthase subunit Atp9 at 37°C. The amount of each mitochondrial genome-encoded protein in mutant cells was determined relative to that in wild-type cells in the experiment described in Fig. 6. (TIF)
Data
The respiratory phenotype of Δubp9 Δubp13 and Δduf1 mutants is not due to a general decrease in free ubiquitin levels. Crude extracts were prepared from cells grown on solid glucose, under the conditions described in Fig. 1 (stationary phase). Extracts from equivalent numbers of cells (based on OD units) were separated in a 5% to 16% MES polyacryla...
Data
Ubp9, Ubp13 and Duf1 display dual localization in soluble and membrane-bound fractions. A. Protoplasts were prepared from cells grown on galactose medium and expressing chromosome-encoded Ubp9-HA (YDB105), Ubp13-HA (YDB106), Duf1-HA (YDB107) or Ubp16-GFP. Aliquots of 13,000 g pellets (P13) and supernatants (S13) corresponding to equivalent numbers...
Data
Duf1 is an unstable, ubiquitylated protein, further destabilized in the absence of its two protein partners, Ubp9 and Ubp13. A. Steady-state levels of Duf1 decrease in the Δubp9 Δubp13 double mutant. Crude extracts prepared from cells expressing chromosome-encoded Duf1-HA in wild-type, Δubp9, Δubp13 and Δubp9 Δubp13 backgrounds were grown on glucos...
Data
Analysis of ATP9 mRNA 5′-end maturation by primer extension. Autoradiographs of 10% polyacrylamide denaturing SDS-PAGE gels on which the products of primer extension were separated. Yeast strains were cultured in the presence of either glucose or lactate as the carbon source (as indicated above the lanes) at two temperatures, 30°C and 37°C. Two dif...
Article
Full-text available
In yeast, the sorting of transmembrane proteins into the multivesicular body (MVB) internal vesicles requires their ubiquitylation by the ubiquitin ligase Rsp5. This allows their recognition by the ubiquitin-binding domains (UBDs) of several endosomal sorting complex required for transport (ESCRT) subunits. K63-linked ubiquitin (K63Ub) chains decor...
Article
Full-text available
For many years, lipid droplets (LDs) were considered to be an inert store of lipids. However, recent data showed that LDs are dynamic organelles playing an important role in storage and mobilization of neutral lipids. In this paper, we report the characterization of LOA1 (alias VPS66, alias YPR139c), a yeast member of the glycerolipid acyltransfera...
Article
Full-text available
Any of seven lysine residues on ubiquitin can serve as the base for chain-extension, resulting in a sizeable spectrum of ubiquitin modifications differing in chain length or linkage type. By optimizing a procedure for rapid lysis, we charted the profile of conjugated cellular ubiquitin directly from whole cell extract. Roughly half of conjugated ub...
Article
Previous studies in the yeast Saccharomyces cerevisiae have proposed a vacuolar localization for Ath1, which is difficult to reconcile with its ability to hydrolyze exogenous trehalose. We used fluorescent microscopy to show that the red fluorescent protein mCherry fused to the C-terminus of Ath1, although mostly localized in the vacuole, was also...
Article
Full-text available
The subcellular localization of plasma membrane proteins, such as receptors and transporters, must be finely tuned so that they can be readily downregulated in response to environmental cues. Some of these membrane proteins are post-translationally modified by conjugation to ubiquitin, which is used as a molecular tag to commit them to the endocyti...
Article
Full-text available
The ubiquitin ligase (E3) Rsp5p is the only member of the Nedd (neural-precursor-cell-expressed, developmentally down-regulated) 4 family of E3s present in yeast. Rsp5p has several proteasome-independent functions in membrane protein trafficking, including a role in the ubiquitination of most plasma membrane proteins, leading to their endocytosis....
Article
Full-text available
The ubiquitylation of membrane proteins destined for the vacuole/lysosome is essential for their recognition by the endosomal sorting machinery and their internalization into vesicles of multivesicular bodies (MVBs). In yeast, this process requires Rsp5p, an essential ubiquitin ligase of the Nedd4 family. We describe here two redundant proteins, Ea...
Article
Peroxisome biogenesis requires the action of about 32 PEX genes encod- ing a family of proteins known as peroxins [1]. These are distributed in the cytosol, peroxisomal membrane, or peroxisome lumen. While a number of these proteins, particularly components of a peroxisome membrane-associated complex known as the importomer [2], regulate the entry...
Article
Full-text available
We identified a cysteine residue, conserved near the N terminus of Pex5p- and Pex20p-like proteins, that is essential for the cytosolic relocation of peroxisomal Pex20p. Surprisingly, this residue is not completely essential for the function of the protein; its point mutation into a serine in Pex20p(C8S) causes the accumulation of the protein at th...
Article
Full-text available
With the approaching completion of the Pichia pastoris genome, a greater emphasis will have to be placed on the proteome and the protein-protein interactions between its constituents. This chapter discusses methods that have been used for the study of such interactions among both soluble and membrane-associated proteins in peroxisome biogenesis. Th...
Article
Full-text available
Based on earlier suggestions that peroxisomes may have arisen from endosymbionts that later lost their DNA, it was expected that protein transport into this organelle would have parallels to systems found in other organelles of endosymbiont origin, such as mitochondria and chloroplasts. This review highlights three features of peroxisomal matrix pr...
Article
Full-text available
Among peroxins involved in peroxisome biogenesis, only Pex8p is predominantly intraperoxisomal at steady state. Pex8p is necessary for peroxisomal matrix protein import via the PTS1 and PTS2 pathways. It is proposed to bridge two peroxisomal membrane subcomplexes comprised of the docking (Pex13p, Pex14p, Pex17p) and RING (Pex2p, Pex10p, Pex12p) per...
Article
Full-text available
We characterize the peroxin PpPex20p from Pichia pastoris and show its requirement for translocation of PTS2 cargoes into peroxisomes. PpPex20p docks at the peroxisomal membrane and translocates into peroxisomes. Its peroxisomal localization requires the docking peroxin Pex14p but not the peroxins Pex2p, Pex10p, and Pex12p, whose absence causes per...
Article
Full-text available
Isu are scaffold proteins involved in iron-sulfur cluster biogenesis and playing a key role in yeast mitochondria and Escherichia coli. In this work, we have characterized the Arabidopsis thaliana Isu gene family. AtIsu1,2,3 genes encode polypeptides closely related to their bacterial and eukaryotic counterparts. AtIsu expression in a Saccharomyces...
Article
Full-text available
Recent results are in favour of a role for NFU-like proteins in Fe-S cluster biogenesis. These polypeptides share a conserved CXXC motif in their NFU domain. In the present study, we have characterized Arabidopsis thaliana NFU1-5 genes. AtNFU proteins are separated into two classes. NFU4 and NFU5 are part of the mitochondrial type, presenting a str...
Article
Full-text available
NifS-like proteins are cysteine desulphurases required for the mobilization of sulphur from cysteine. They are present in all organisms, where they are involved in iron-sulphur (Fe-S) cluster biosynthesis. In eukaryotes, these enzymes are present in mitochondria, which are the major site for Fe-S cluster assembly. The genome of the model plant Arab...
Article
Full-text available
Under phosphorous deficiency, plants of white lupin (Lupinus albus L.) develop root clusters, which are also called proteoid roots due to their preferential presence in the Proteaceae. In their mature stage, these roots acidify the soil and excrete high amounts of carboxylates [up to 1.5 and 7 mol (g FW)-1 h-1 of malate and citrate, respectively] e...

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Projects (4)
Project
Cancer cells increase their glucose intake in support of their proliferative metabolism based on aerobic glycolysis (Warburg effect). Pharmacological targeting of glycolysis has been proposed to treat cancer, including the use of 2-deoxyglucose (2DG), a metabolic inhibitor, combined with other molecules. 2DG is imported into tumor cells and after import, it is phosphorylated in 2DG6P and inhibits glycolysis; it also interferes with glycosylation of proteins, inducing ER stress and then death by apoptosis. However, its cellular effects are not fully understood. We propose to use yeast as a tool to define the molecular basis of 2DG toxicity as well as associated resistance mechanisms through genetics, cell biology and proteomics approaches. This work is funded by the Fondation ARC pour la recherche sur le cancer (PJA20181208080), a post-doctoral fellowship by the Fondation pour la Recherche Médicale (SPF20150934065) and a PhD fellowship by the Ligue contre le cancer (TAZK20115).
Project
We are currently interested in understanding the molecular details of ART regulation by signaling and cell physiology.
Project
Yeast cells rapidly adapt to changes in the nutrients provided in the growth medium. This involves a rewiring of the transcriptional program as well as a reconfiguration of the proteome, such as metabolic enzymes or nutrient transporters. This project focuses on understanding how nutrient transporters homeostasis is regulated by glucose availability, and the contribution of signalling pathways (notably, AMPK/Snf1), arrestins-related proteins and ubiquitin.