Alain Van Dorsselaer

Institut Pluridisciplinaire Hubert Curien, Strasburg, Alsace, France

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Publications (475)2061.48 Total impact

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    ABSTRACT: This study aimed to identify a protein marker that can differentiate between fresh skinless and frozen-thawed sea bass (Dicentrarchus labrax) fillets using the two-dimensional polyacrylamide gel electrophoresis (2-DE) technique. Distinct gel patterns, due to proteins with low molecular weight and low isoelectric points, distinguished fresh fillets from frozen-thawed ones. Frozen-thawed fillets showed two specific protein spots as early as the first day of the study. However, these spots were not observed in fresh fillets until at least 13 days of storage between 0 and 4°C, fillets were judged, beyond this period, fish were unfit for human consumption as revealed by complementary studies on fish spoilage indicators namely total volatile basic nitrogen and biogenic amines. Mass spectrometry identified the specific proteins as parvalbumin isoforms. Parvalbumins may thus be useful markers of differentiation between fresh and frozen-thawed sea bass fillets.
    Food Chemistry 06/2015; 176:294-301. DOI:10.1016/j.foodchem.2014.12.065 · 3.26 Impact Factor
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    ABSTRACT: The bioproduction of recombinant monoclonal antibodies (mAbs) results in complex mixtures of a main isoform and numerous macro and micro-variants. MAbs present therefore different levels of heterogeneities ranging from primary sequence variants to post-translational modifications. Among these heterogeneities are truncation and fragmentation of the primary amino-acid sequence resulting in shorter or cleaved polypeptide chains. Another reported heterogeneity comes from the incomplete processing of the signal peptide resulting in N-terminal elongated polypeptide chains. Here we present a novel in-gel protein N-terminal chemical derivatization method with trimethoxyphenyl phosphonium (TMPP) that enhances the mass spectrometry detection of N-terminal positions of proteins and allows their unambiguous assignment without altering the identification of internal digestion peptides. This method requires only one additional step to a classical peptide mapping workflow. This in-gel N-TOP (N-Terminal Oriented Proteomics) strategy was applied successfully to detect and characterize the N-terminal sequence heterogeneities of several monoclonal antibodies among which minor unexpected proteoforms.
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    ABSTRACT: Antibody-drug conjugates (ADCs) are bio-chemotherapeutics consisting of a cytotoxic chemical drug linked covalently to a monoclonal antibody. Two main classes of ADCs, namely cysteine and lysine conjugates, are currently available on the market or involved in clinical trials. The complex structure and heterogeneity of ADCs makes their biophysical characterization challenging. For cysteine conjugates, hydrophobic interaction chromatography is the gold standard technique for studying drug distribution, the naked antibody content, and the average drug to antibody ratio (DAR). For lysine ADC conjugates on the other hand, which are not amenable to hydrophobic interaction chromatography because of their higher heterogeneity, denaturing mass spectrometry (MS) and UV/Vis spectroscopy are the most powerful approaches. We report here the use of native MS and ion mobility (IM-MS) for the characterization of trastuzumab emtansine (T-DM1, Kadcyla®). This lysine conjugate is currently being considered for the treatment of human epidermal growth factor receptor 2 (HER2)-positive breast cancer, and combines the anti-HER2 antibody trastuzumab (Herceptin®), with the cytotoxic microtubule-inhibiting maytansine derivative, DM1. We show that native MS combined with high-resolution measurements and/or charge reduction is beneficial in terms of the accurate values it provides of the average DAR and the drug load profiles. The use of spectral deconvolution is discussed in detail. We report furthermore the use of native IM-MS to directly determine DAR distribution profiles and average DAR values, as well as a molecular modeling investigation of positional isomers in T-DM1. This article is protected by copyright. All rights reserved. © 2015 The Protein Society.
    Protein Science 02/2015; DOI:10.1002/pro.2666 · 2.86 Impact Factor
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    ABSTRACT: The approval process for antibody biosimilars relies primarily on comprehensive analytical data to establish comparability and high similarity with the originator. Mass spectrometry (MS) in combination with liquid chromatography (LC) and electrophoretic methods are the corner stone for comparability and biosimilarity evaluation. In this special feature we report head-to-head comparison of trastuzumab and cetuximab with corresponding biosimilar and biobetter candidates based on cutting-edge mass spectrometry techniques such as native MS and ion-mobility MS at different levels (top, middle and bottom). In addition, we discuss the advantages and the limitations of sample preparation and enzymatic digestion, middle-up and -down strategies and the use of hydrogen/deuterium exchange followed by MS (HDX-MS). Last but not least, emerging separation methods combined to MS such as capillary zone electrophoresis-tandem MS (CESI-MS/MS), electron transfer dissociation (ETD), top down-sequencing (TDS) and high-resolution MS (HR-MS) that complete the panel of state-of-the-art MS-based options for comparability and biosimilarity evaluation are presented. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.
    Journal of Mass Spectrometry 02/2015; 50(2). DOI:10.1002/jms.3554 · 2.71 Impact Factor
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    ABSTRACT: Site-specific isomerization of uridines into pseudouridines in RNAs is catalyzed either by stand-alone enzymes or by box H/ACA ribonucleoprotein particles (sno/sRNPs). The archaeal box H/ACA sRNPs are 5-component complexes that consist of a guide RNA and the aCBF5, aNOP10, L7Ae and aGAR1 proteins. In this study, we performed pairwise incubations of individual constituents of archaeal box H/ACA sRNPs and analyzed their interactions by native mass spectrometry (MS) to build a 2D connectivity map of direct binders. We describe the use of native MS in combination with ion mobility-MS to monitor the in vitro assembly of the active H/ACA sRNP particle. Real-time native MS was used to monitor how box H/ACA particle functions in multiple-turnover conditions. Native MS also unambiguously revealed that a substrate RNA containing 5-fluorouridine (f(5) U) was hydrolyzed into 5-fluoro-6-hydroxy-pseudouridine (f(5) ho(6) Ψ). In terms of enzymatic mechanism, box H/ACA sRNP was shown to catalyze the pseudouridylation of a first RNA substrate, then to release the RNA product (S22 f(5) ho(6) ψ) from the RNP enzyme and reload a new substrate RNA molecule. Altogether, our native MS-based approaches provide relevant new information about the potential assembly process and catalytic mechanism of box H/ACA RNPs. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Proteomics 02/2015; DOI:10.1002/pmic.201400529 · 3.97 Impact Factor
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    ABSTRACT: Amine-to-thiol coupling is the most common route for the preparation of antibody-drug conjugates (ADC). It is usually achieved by using heterobifunctional reagents possessing an activated ester at one end and a maleimide group at the other. However, maleimide-based conjugates were recently revealed to have limited stability in blood circulation, which can compromise therapeutic efficacy of the conjugate. To address this issue, we have developed a heterobifunctional reagent, sodium 4-((4-(cyanoethynyl)benzoyl)oxy)-2,3,5,6-tetrafluorobenzenesulfonate (CBTF), for amine-to-thiol coupling. It comprises a recently described 3-arylpropionitrile (APN) function in replacement of maleimide and allows for the preparation of remarkably stable conjugates. A series of antibody-dye conjugates have been prepared using this reagent and shown superior stability in human blood plasma compared to maleimide-derived conjugates.
    Bioconjugate Chemistry 01/2015; 26(2). DOI:10.1021/bc500610g · 4.82 Impact Factor
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    ABSTRACT: Titanium dioxide and copper oxide nanoparticles are more and more widely used because of their catalytic properties, of their light absorbing properties (titanium dioxide) or of their biocidal properties (copper oxide), increasing the risk of adverse health effects. In this frame, the responses of mouse macrophages were studied. Both proteomic and targeted analyses were performed to investigate several parameters, such as phagocytic capacity, cytokine release, copper release, and response at sub toxic doses. Besides titanium dioxide and copper oxide nanoparticles, copper ions were used as controls. We also showed that the overall copper release in the cell does not explain per se the toxicity observed with copper oxide nanoparticles. In addition, both copper ion and copper oxide nanoparticles, but not titanium oxide, induced DNA strands breaks in macrophages. As to functional responses, the phagocytic capacity was not hampered by any of the treatments at non-toxic doses, while copper ion decreased the lipopolysaccharide-induced cytokine and nitric oxide productions. The proteomic analyses highlighted very few changes induced by titanium dioxide nanoparticles, but an induction of heme oxygenase, an increase of glutathione synthesis and a decrease of tetrahydrobiopterin in response to copper oxide nanoparticles. Subsequent targeted analyses demonstrated that the increase in glutathione biosynthesis and the induction of heme oxygenase (e.g. by lovastatin/monacolin K) are critical for macrophages to survive a copper challenge, and that the intermediates of the catecholamine pathway induce a strong cross toxicity with copper oxide nanoparticles and copper ions.
    PLoS ONE 01/2015; 10(4):e0124496. DOI:10.1371/journal.pone.0124496 · 3.53 Impact Factor
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    ABSTRACT: To assess a new adenovirus-based immunotherapy as a novel treatment approach to chronic hepatitis B (CHB). TG1050 is a non-replicative adenovirus serotype 5 encoding a unique large fusion protein composed of a truncated HBV Core, a modified HBV Polymerase and two HBV Envelope domains. We used a recently described HBV-persistent mouse model based on a recombinant adenovirus-associated virus encoding an over length genome of HBV that induces the chronic production of HBsAg, HBeAg and infectious HBV particles to assess the ability of TG1050 to induce functional T cells in face of a chronic status. In in vitro studies, TG1050 was shown to express the expected large polyprotein together with a dominant, smaller by-product. Following a single administration in mice, TG1050 induced robust, multispecific and long-lasting HBV-specific T cells detectable up to 1 year post-injection. These cells target all three encoded immunogens and display bifunctionality (ie, capacity to produce both interferon γ and tumour necrosis factor α as well as cytolytic functions). In addition, control of circulating levels of HBV DNA and HBsAg was observed while alanine aminotransferase levels remain in the normal range. Injection of TG1050 induced both splenic and intrahepatic functional T cells producing cytokines and displaying cytolytic activity in HBV-naïve and HBV-persistent mouse models together with significant reduction of circulating viral parameters. These results warrant clinical evaluation of TG1050 in the treatment of CHB. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.
    Gut 11/2014; DOI:10.1136/gutjnl-2014-308041 · 13.32 Impact Factor
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    ABSTRACT: Antibody drug conjugates (ADCs) are macromolecules composed of cytotoxic drugs covalently attached via a conditionally stable linker to monoclonal antibodies (mAbs). ADCs are among most promising next generation of empowered mAbs foreseen to treat cancers. Compared to naked mAbs, ADCs have increased level of complexity as the heterogeneity of conjugation cumulates with the inherent micro-variability of the biomolecule. An increasing need underlying ADC's development and optimization is to improve its analytical and bioanalytical characterization by assessing three main ADC quality attributes: drug distribution, amount of naked antibody and average drug to antibody ratio (DAR). Here, the analytical potential of native mass spectrometry (MS) and native ion mobility MS (IM-MS) is compared to hydrophobic interaction chromatography (HIC), the reference method for quality control of interchain cysteinyl-linked ADCs. Brentuximab vedotin, first in class and gold standard, was chosen for a proof of principle. High resolution native MS provided accurate mass measurement (<30 ppm) of intact ADCs together with average DAR and drug distribution, confirming the unique ability of native MS for simultaneous detection of mixtures of covalent and non-covalent products. Native IM-MS was next used for the first time to characterize an ADC. IM-MS evidenced ADC multiple drug loading, collisional cross sections measurement of each payload species attesting slight conformational changes. A semi-quantitative interpretation of IM-MS data was developed to directly extrapolate average DAR and DAR distribution. Additionnaly, HIC fractions were collected and analyzed by native MS and IM-MS, assessing the interpretation of each HIC peak. Altogether, our results illustrate how native MS and IM-MS can rapidly assess ADC structural heterogeneity and how easily these methods can be implemented into MS workflows for in-depth ADC analytical characterization.
    Analytical Chemistry 09/2014; 86(21). DOI:10.1021/ac502593n · 5.83 Impact Factor
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    ABSTRACT: Biogenesis of eukaryotic box C/D small nucleolar ribonucleoprotein particles (C/D snoRNPs) involves conserved trans-acting factors, which are proposed to facilitate the assembly of the core proteins Snu13p/15.5K, Nop58p/NOP58, Nop56p/NOP56 and Nop1p/Fibrillarin on box C/D small nucleolar RNAs (C/D snoRNAs). In yeast, protein Rsa1 acts as a platform, interacting with both the RNA-binding core protein Snu13 and protein Pih1 of the Hsp82-R2TP chaperone complex. In this work, a proteomic approach coupled with functional and structural studies identifies protein Hit1 as a novel Rsa1p-interacting partner involved in C/D snoRNP assembly. Hit1p contributes to in vivo C/D snoRNA stability and pre-RNA maturation kinetics. It associates with U3 snoRNA precursors and influences its 3'-end processing. Remarkably, Hit1p is required to maintain steady-state levels of Rsa1p. This stabilizing activity is likely to be general across eukaryotic species, as the human protein ZNHIT3(TRIP3) showing sequence homology with Hit1p regulates the abundance of NUFIP1, the Rsa1p functional homolog. The nuclear magnetic resonance solution structure of the Rsa1p317-352-Hit1p70-164 complex reveals a novel mode of protein-protein association explaining the strong stability of the Rsa1p-Hit1p complex. Our biochemical data show that C/D snoRNAs and the core protein Nop58 can interact with the purified Snu13p-Rsa1p-Hit1p heterotrimer.
    Nucleic Acids Research 08/2014; 42(16). DOI:10.1093/nar/gku612 · 8.81 Impact Factor
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    ABSTRACT: Complete starvation may prove lethal due to excessive loss of body proteins. However, it is still not completely understood whether responses to food deprivation are time-dependently induced or triggered in relation with the successive phases of protein sparing and wasting that characterize prolonged fasting. As the liver has a wide range of vital functions, we examined the hepatic regulatory mechanisms elicited during prolonged fasting. We showed that fasting-induced transcriptome/proteome changes occur in close relation with fuel partitioning, independently of ATP levels. Omics data suggesting a worsening of oxidative stress during the proteolytic stage of fasting, this was further validated using biochemical assays. Low levels of antioxidant factors were indeed paralleled by their decreased activity, which could be impaired by low NADPH levels. Oxidative damages on lipids and proteins were accordingly increased only during late fasting. At this stage, the gene/protein expression of several chaperones was also repressed. Together with the impairment of metabolic achievements, a vicious cycle involving protein misfolding and oxidative stress could jeopardize liver functions when the proteolytic stage of fasting is reached. Thus, monitoring of liver impairments should help to better manage or treat catabolic and/or oxidative stress conditions, such as ageing and degeneration. This article is protected by copyright. All rights reserved.
    Proteomics 08/2014; 14(16). DOI:10.1002/pmic.201400051 · 3.97 Impact Factor
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    ABSTRACT: Botryosphaeria dieback is a fungal grapevine trunk disease that currently represents a threat for viticulture worldwide because of the important economical losses due to reduced yield of affected plants and their premature death. Neofusicoccum parvum and Diplodia seriata are among the causal agents. Vine green stems were artificially infected with N. parvum or D. seriata at the onset of three different phenological stages (G stage (separated clusters), flowering and veraison). Highest mean lesion lengths were recorded at flowering. Major proteome changes associated to artificial infections during the three different phenological stages were also reported using two dimensional gel electrophoresis (2D)-based analysis. Twenty (G stage), 15 (flowering) and 13 (veraison) differentially expressed protein spots were subjected to nanoLC-MS/MS and a total of 247, OPEN ACCESS Int. J. Mol. Sci. 2014, 15 9645 54 and 25 proteins were respectively identified. At flowering, a weaker response to the infection was likely activated as compared to the other stages, and some defense-related proteins were even down regulated (e.g., superoxide dismutase, major latex-like protein, and pathogenesis related protein 10). Globally, the flowering period seemed to represent the period of highest sensitivity of grapevine to Botryosphaeria dieback agent infection, possibly being related to the high metabolic activity in the inflorescences.
    International Journal of Molecular Sciences 05/2014; 15(6):9644-9669. DOI:10.3390/ijms15069644 · 2.34 Impact Factor
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    ABSTRACT: Background Life history theories predict that investment in current reproduction comes at a cost for future reproduction and survival. Oxidative stress is one of the best documented mechanisms underlying costs of reproduction to date. However, other, yet to be described molecular mechanisms that play a short term role during reproduction may explain the negative relationships underlying the cost of reproduction. To identify such new mechanisms, we used a global proteomic determination of liver protein profiles in laboratory adult female mice whose litter size had been either reduced or enlarged after birth. This litter size manipulation was expected to affect females by either raising or decreasing their current reproductive effort. At the same time, global parameters and levels of oxidative stress were also measured in all females. Results Based on plasma analyses, females with enlarged litters exhibited increased levels of oxidative stress at the date of weaning compared to females with reduced litters, while no significant difference was found between both the latter groups and control females. None of the liver proteins related to oxidative balance were significantly affected by the experimental treatment. In contrast, the liver protein profiles of females with enlarged and reduced litters suggested that calcium metabolism and cell growth regulation were negatively affected by changes in the number of pup reared. Conclusions Plasma oxidative stress levels in reproductive mice revealed that the degree of investment in reproduction can actually incur a cost in terms of plasmatic oxidative stress, their initial investment in reproduction being close to maximum and remaining at a same level when the energy demand of lactation is reduced. Liver proteomic profiles in reproductive females show that hepatic oxidative stress is unlikely to be involved in the cost of reproduction. Reproductive females rather exhibited liver protein profiles similar to those previously described in laboratory ageing mice, thus suggesting that hepatic cell pro-ageing processes may be involved in the cost of reproduction. Overall, our data illustrate how a proteomic approach can unravel new mechanisms sustaining life-history trade-offs, and reproduction costs in particular.
    Frontiers in Zoology 05/2014; 11:41. DOI:10.1186/1742-9994-11-41 · 2.30 Impact Factor
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    ABSTRACT: Among fungal grapevine trunk diseases, esca proper poses a significant threat for viticulture. Apoplexy, mainly occurring on grapevines affected by esca proper, is also a threat. To verify if different responses are activated in the woody tissues of apoplectic (A) and esca proper-affected (E) vines, two-dimensional gel electrophoresis coupled to mass spectrometry analysis was used to examine changes in the trunk wood of E and A field-grown plants. Asymptomatic and black streaked trunk (BST) wood from A and E plants were compared to asymptomatic and BST wood of control plants. Twenty-seven differentially expressed protein spots were identified. For eleven targeted proteins, expression of the relative transcripts was also monitored by qRT-PCR. Hierarchical tree clustering revealed differences in the distribution of spots containing carbohydrate metabolism proteins and heat shock proteins between asymptomatic- and BST-wood of grapevine, irrespective of the type of plant examined (control or diseased grapevines). Asymptomatic wood was mainly characterized by down-expression of proteins involved in cell growth and defense responses. The proteome of BST wood, characterized by extensive presence of grapevine trunk disease agents, revealed over-expression of proteins involved in defense. There was no evidence of strong different response in the trunk wood of apoplectic and esca proper affected plants. This could mean that, despite the different feature of these external crown symptoms, grapevine responses at trunk wood level is very similar in both cases. This plant response might therefore either simply be due to the fact that plants can react in the same way to different stresses, or that apoplexy represents a different effect provoked by the same cause or causal agent(s).
    Phytopathologia Mediterranea 05/2014; 53(1):168-187. DOI:10.14601/Phytopathol_Mediterr-12913 · 1.29 Impact Factor
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    ABSTRACT: Fusarium graminearum was grown on four lignocellulosic substrates (corn cobs, wheat bran, hop cell walls, and birchwood) and glucose as the sole carbon source. Proteomic studies performed on the resulting enzymatic cocktails highlighted a great diversity in the number and type of proteins secreted. The cell wall degrading enzymes (CWDE) proportion varied greatly from 20% to 69%. Only 1 of the 57 CWDEs detected in this study was common to the five proteomes. In contrast, 35 CWDEs were specific to one proteome only. The polysaccharide-degradation activities were different depending on the cocktail and the polysaccharide used. F. graminearum strongly modifies the enzymatic cocktail it secretes as a function of the biomass used for growth. This article is protected by copyright. All rights reserved.
    FEMS Microbiology Letters 05/2014; 355(2). DOI:10.1111/1574-6968.12467 · 2.72 Impact Factor
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    ABSTRACT: One of the major bottlenecks in the proteomics field today resides in the computational interpretation of the massive data generated by the latest generation of high-throughput mass spectrometry instruments. MS/MS datasets are constantly increasing in size and complexity and it becomes challenging to comprehensively process such huge datasets and afterwards deduce most relevant biological information. The Mass Spectrometry Data Analysis (MSDA, https://msda.unistra.fr) online software suite provides a series of modules for in-depth MS/MS data analysis. It includes a custom databases generation toolbox, modules for filtering and extracting high-quality spectra, for running high performance database and de novo searches, and for extracting modified peptides spectra and functional annotations. Additionally, MSDA enables running the most computationally intensive steps, i.e. database and de novo searches, on a computer grid thus providing a net time gain of up to 99% for data processing. This article is protected by copyright. All rights reserved.
    Proteomics 05/2014; 14(9). DOI:10.1002/pmic.201300415 · 3.97 Impact Factor
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    ABSTRACT: Two different zinc oxide nanoparticles, as well as zinc ions, are used to study the cellular responses of the RAW 264 macrophage cell line. A proteomic screen is used to provide a wide view of the molecular effects of zinc, and the most prominent results are cross-validated by targeted studies. Furthermore, the alteration of important macrophage functions (e.g. phagocytosis) by zinc is also investigated. The intracellular dissolution/uptake of zinc is also studied to further characterize zinc toxicity. Zinc oxide nanoparticles dissolve readily in the cells, leading to high intracellular zinc concentrations, mostly as protein-bound zinc. The proteomic screen reveals a rather weak response in the oxidative stress response pathway, but a strong response both in the central metabolism and in the proteasomal protein degradation pathway. Targeted experiments confirm that carbohydrate catabolism and proteasome are critical determinants of sensitivity to zinc, which also induces DNA damage. Conversely, glutathione levels and phagocytosis appear unaffected at moderately toxic zinc concentrations.
    Nanoscale 05/2014; 6(11). DOI:10.1039/c4nr00319e · 6.74 Impact Factor
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    ABSTRACT: Botryosphaeria dieback is a fungal grapevine trunk disease which represents a threat for viticulture worldwide due to the decreased production of affected plants and their premature death. This dieback is characterized by a typical wood discoloration called "brown stripe". Herein, a proteome comparison of the brown striped wood from Botryosphaeria dieback-affected standing vines cultivar 'Chardonnay', 'Gewurztraminer' and 'Mourvèdre' was performed. The transcript analysis for 15 targeted genes and the quantification of both total phenolics and specific stilbenes were also performed. Several pathogenesis-related proteins and members of the antioxidant system were more abundant in the brown striped wood of the three cultivars, whereas other defense-related proteins were less abundant. Additionally, total phenolics and some specific stilbenes were more accumulated in the brown striped wood. Strongest differences among the cultivars concerned especially proteins of the primary metabolism, which looked to be particularly impaired in the brown striped wood of 'Chardonnay'. Low abundance of some proteins involved in defense response probably contributes to make global response insufficient to avoid the symptom development. The differential susceptibility of the three grapevine cultivars could be linked to the diverse expression of various proteins involved in defense response, stress tolerance and metabolism.
    Phytopathology 04/2014; 104(10). DOI:10.1094/PHYTO-01-14-0007-R · 2.75 Impact Factor
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    ABSTRACT: Glioblastomas (GBMs) are highly aggressive, invasive brain tumors with bad prognosis and unmet medical need. These tumors are heterogeneous being constituted by a variety of cells in different states of differentiation. Among these, cells endowed with stem properties, tumor initiating/propagating properties and particularly resistant to chemo- and radiotherapies are designed as the real culprits for tumor maintenance and relapse after treatment. These cells, termed cancer stem-like cells, have been designed as prominent targets for new and more efficient cancer therapies. G-protein coupled receptors (GPCRs), a family of membrane receptors, play a prominent role in cell signaling, cell communication and crosstalk with the microenvironment. Their role in cancer has been highlighted but remains largely unexplored. Here, we report a descriptive study of the differential expression of the endo-GPCR repertoire in human glioblastoma cancer stem-like cells (GSCs), U-87 MG cells, human astrocytes and fetal neural stem cells (f-NSCs). The endo-GPCR transcriptome has been studied using Taqman Low Density Arrays. Of the 356 GPCRs investigated, 138 were retained for comparative studies between the different cell types. At the transcriptomic level, eight GPCRs were specifically expressed/overexpressed in GSCs. Seventeen GPCRs appeared specifically expressed in cells with stem properties (GSCs and f-NSCs). Results of GPCR expression at the protein level using mass spectrometry and proteomic analysis are also presented. The comparative GPCR expression study presented here gives clues for new pathways specifically used by GSCs and unveils novel potential therapeutic targets.
    PLoS ONE 03/2014; 9(3):e91519. DOI:10.1371/journal.pone.0091519 · 3.53 Impact Factor
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    ABSTRACT: The nuclear bile acid receptor farnesoid X receptor (FXR) is an important transcriptional regulator of bile acid, lipid, and glucose metabolism. FXR is highly expressed in the liver and intestine and controls the synthesis and enterohepatic circulation of bile acids. However, little is known about FXR-associated proteins that contribute to metabolic regulation. Here, we performed a mass spectrometry-based search for FXR-interacting proteins in human hepatoma cells and identified AMPK as a coregulator of FXR. FXR interacted with the nutrient-sensitive kinase AMPK in the cytoplasm of target cells and was phosphorylated in its hinge domain. In cultured human and murine hepatocytes and enterocytes, pharmacological activation of AMPK inhibited FXR transcriptional activity and prevented FXR coactivator recruitment to promoters of FXR-regulated genes. Furthermore, treatment with AMPK activators, including the antidiabetic biguanide metformin, inhibited FXR agonist induction of FXR target genes in mouse liver and intestine. In a mouse model of intrahepatic cholestasis, metformin treatment induced FXR phosphorylation, perturbed bile acid homeostasis, and worsened liver injury. Together, our data indicate that AMPK directly phosphorylates and regulates FXR transcriptional activity to precipitate liver injury under conditions favoring cholestasis.
    The Journal of clinical investigation 02/2014; 124(3). DOI:10.1172/JCI68815 · 13.77 Impact Factor

Publication Stats

13k Citations
2,061.48 Total Impact Points

Institutions

  • 2007–2014
    • Institut Pluridisciplinaire Hubert Curien
      • Department of Ecology, Physiology and Ethology
      Strasburg, Alsace, France
    • University of Leipzig
      • Institut für Biochemie
      Leipzig, Saxony, Germany
  • 1985–2014
    • University of Strasbourg
      • Laboratoire de Chimie Organique des Substances Naturelles
      Strasburg, Alsace, France
  • 1983–2014
    • French National Centre for Scientific Research
      • • Institut des maladies neurodégénératives (MN)
      • • Centre d'Ecologie Fonctionnelle et Evolutive
      • • Centre de Biochimie Structurale
      • • Institute for Molecular and Cellular Biology (IBMC)
      Lutetia Parisorum, Île-de-France, France
  • 2013
    • University of Grenoble
      Grenoble, Rhône-Alpes, France
    • Institut Pasteur de Lille
      Lille, Nord-Pas-de-Calais, France
  • 2010
    • NovAliX
      Illkirch, Alsace, France
  • 2002–2010
    • Universidad de La Punta
      La Punta, San Luis, Argentina
    • University of Bologna
      • "Giacomo Ciamician" Department of Chemistry CHIM
      Bolonia, Emilia-Romagna, Italy
    • Université Libre de Bruxelles
      • Unit of Organic and Physical Chemistry
      Bruxelles, Brussels Capital, Belgium
  • 2009
    • University Joseph Fourier - Grenoble 1
      • Institut Albert Bonniot
      Grenoble, Rhone-Alpes, France
  • 2008
    • National Institute of Livestock and Grassland Science
      Ibaragi, Ōsaka, Japan
    • Claude Bernard University Lyon 1
      Villeurbanne, Rhône-Alpes, France
  • 2002–2007
    • University of Waterloo
      • Department of Chemistry
      Waterloo, Ontario, Canada
  • 2006
    • Universität des Saarlandes
      Saarbrücken, Saarland, Germany
    • State University of New York College at Old Westbury
      Old Westbury, New York, United States
  • 2005
    • Institut de Génétique et de Biologie Moléculaire et Cellulaire
      Strasburg, Alsace, France
  • 2004
    • Institut de Physique et Chimie des Matériaux de Strasbourg
      Strasburg, Alsace, France
  • 1997
    • French Institute of Health and Medical Research
      Lutetia Parisorum, Île-de-France, France
    • University of Geneva
      • Department of Biochemistry
      Genève, Geneva, Switzerland
  • 1995
    • Unité Inserm U1077
      Caen, Lower Normandy, France
  • 1994
    • European Molecular Biology Laboratory
      Heidelburg, Baden-Württemberg, Germany
  • 1992–1994
    • Universität Basel
      • Department of Biophysical Chemistry
      Basel, BS, Switzerland
  • 1987–1992
    • Natural Product Chemistry Institute
      Lutetia Parisorum, Île-de-France, France
  • 1991
    • Odense University Hospital
      • Molecular biology laboratory
      Odense, South Denmark, Denmark
  • 1989
    • Transgene
      Illkirch, Alsace, France