Éditeur(s) : HAL CCSD
Résumé : Scope: TNF-Related Apoptosis-Inducing Ligand (TRAIL) induces apoptosis via the extrinsic death pathway in human colon cancer cells. Short Chain Fatty Acids (SCFA) by contrast induce apoptosis of cancer cells via the intrinsic death pathway. We have previously shown that food-grade dairy propionibacteria induce intrinsic apoptosis of colon cancer cells, via SCFA metabolites (propionate and acetate) acting on mitochondria. We investigate here the possible synergistic pro-apoptotic effect between Propionibacterium freudenreichii and TRAIL. Methods and results: Whole transcriptomic analysis demonstrated that propionibacterial supernatant or propionibacterial SCFA, in combination with TRAIL, increased pro-apoptotic gene expression (TRAIL-R2/DR5) and decreased anti-apoptotic gene expression (FLIPL, XIAP) in HT29 human colon cancer cells. The revealed synergistic pro-apoptotic effect, depending on both death receptors (TRAIL-R1/DR4, TRAIL-R2/DR5) and caspases (caspase-8, -9 and -3) activation, was lethal on cancer cells but not on normal human intestinal epithelial cells (HIEC), and was inhibited by Bcl-2 expression. Finally, milk fermented by P. freudenreichii induced HT29 cells apoptosis and enhanced TRAIL cytotoxic activity, as did P. freudenreichii culture supernatants or its SCFA metabolites.Conclusion: The revealed synergy opens new applications for food-grade P. freudenreichii-containing products as new adjuvants to potentiate TRAIL-based cancer therapy in colorectal cancer.
International Scientific Conference on Probiotics and Prebiotics - IPC2016
Budapest, Hungary

hal-01286497
https://hal.archives-ouvertes.fr/hal-01286497
PRODINRA : 349367

Éditeur(s) : HAL CCSD American Society for Investigative Pathology
Résumé : International audience
Based on both experimental and clinical data, tamoxifen has been proposed to have cardiovascular benefits, although the mechanism(s) contributing to that protective effect are still poorly understood. In vitro experiments demonstrated that tamoxifen elicits its transcriptional effect through estrogen receptor (ER) α, but other targets can participate in its actions. However, although tamoxifen selectively activates the activating function (AF)-1 of ERα, we recently showed that this ERα subfunction is dispensable for the atheroprotective action of 17β-estradiol (E2), the main ligand of ERs. The goal of the present work is to determine to which extent ERα and its AF-1 mediate the vasculoprotective action of tamoxifen. Our data confirm that tamoxifen exerts an atheroprotective action on low density lipoprotein receptor (LDL-r(-/-)) female mice, but, in contrast to E2, it fails to accelerate reendothelialization after carotid electric injury. Tamoxifen and E2 elicit differences in gene expression profiles in the mouse aorta. Finally, the atheroprotective action of tamoxifen is abrogated in ERα(-/-)LDL-r(-/-) mice and in LDL-r(-/-) mice selectively deficient in ERαAF-1 (ERαAF-1(0/0)LDL-r(-/-)). Our results demonstrate, for the first time to our knowledge, that tamoxifen mediates its actions in vivo through the selective activation of ERαAF-1, which is sufficient to prevent atheroma, but not to accelerate endothelial healing.
ISSN: 0002-9440

hal-00874317
https://hal.archives-ouvertes.fr/hal-00874317
DOI : 10.1016/j.ajpath.2013.03.010
PUBMED : 23669343

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
: Zebrafish, Danio rerio, is increasingly used as an animal model to study the effects of pharmaceuticals and environmental estrogens. As most of these estrogens have only been tested on human estrogen receptors (ERs), it is necessary to measure their effects on zebrafish ERs. In humans there are two distinct nuclear ERs (hERα and hERβ), whereas the zebrafish genome encodes three ERs, zfERα and two zfERβs (zfERβ1 and zfERβ2). In this study, we established HeLa-based reporter cell lines stably expressing each of the three zfERs. We first reported that estrogens more efficiently activate the zfERs at 28°C as compared to 37°C, thus reflecting the physiological temperature of zebrafish in wildlife. We then showed significant differences in the ability of agonist and antagonist estrogens to modulate activation of the three zfER isotypes in comparison to hERs. Environmental compounds (bisphenol A, alkylphenols, mycoestrogens) which are hER panagonists and hERβ selective agonists displayed greater potency for zfERα as compared to zfERβs. Among hERα selective synthetic agonists, PPT did not activate zfERα while 16α-LE2 was the most zfERα selective compound. Altogether, these results confirm that all hER ligands control in a similar manner the transcriptional activity of zfERs although significant differences in selectivity were observed among subtypes. The zfER subtype selective ligands that we identified thus represent new valuable tools to dissect the physiological roles of the different zfERs. Finally, our work also points out that care has to be taken in transposing the results obtained using the zebrafish as a model for human physiopathology.
ISSN: 0041-008X

inserm-01067804
http://www.hal.inserm.fr/inserm-01067804
http://www.hal.inserm.fr/inserm-01067804/document
http://www.hal.inserm.fr/inserm-01067804/file/pintot_et_al._manuscript.pdf
DOI : 10.1016/j.taap.2014.07.020

Éditeur(s) : HAL CCSD Public Library of Science
Résumé : International audience
Many studies have reported proliferative, differentiating or protective effects of estradiol, notably through estrogen receptor alpha (ERα). On the contrary, the ligand-independent action of ERα is currently poorly documented notably in cell protection. The stable transfection of wild type, substituted or truncated form of ERα in PC12 cells (ERα negative cell line) lead the specific study of its ligand-independent action. Hence, we demonstrate here that, in the absence of E2, the expression of ERα prevents cells from apoptosis induced by serum deprivation. This protection is not due to an ERE-mediated transcription and does not require either AF-1 or AF-2 transactivation functions. It is afforded to the Y537 residue of ERα and activation of c-Src/Stat3 signaling pathway.
ISSN: 1932-6203

inserm-00865332
http://www.hal.inserm.fr/inserm-00865332
http://www.hal.inserm.fr/inserm-00865332/document
http://www.hal.inserm.fr/inserm-00865332/file/PMID_23825704.pdf
DOI : 10.1371/journal.pone.0069081

Éditeur(s) : HAL CCSD
Résumé : International audience
OBJECTIVES: Semen composition is influenced by HIV-1 infection, yet the impact of semen components on HIV infection of primary target cells has only been studied in samples from HIV-uninfected donors. DESIGN: We compared the effect of seminal plasma from chronically HIV-infected (SP+) versus uninfected donors (SP-) on HIV-1 infection of PBMCs and CD4+T cells. METHODS: Primary cells were infected with HIV-1 in the presence of SP+ or SP- and analyzed for infection level, metabolic activity, HIV receptor expression, proliferation and activation. SP+ and SP- were compared for infection-enhancing peptides, cytokines and prostaglandin E2 levels. RESULTS: SP- efficiently enhanced HIV-1 R5 infection of CD4+ T cells, whereas SP+ enhancing activity was significantly reduced. RANTES concentrations were elevated in SP+ relative to SP-, while the concentrations of infectivity-enhancing peptides (SEVI, SEM1, SEM2) were similar. CCR5 membrane expression levels were reduced on CD4+T cells shortly post-exposure to SP+ compared with SP- and correlated to R5-tropic HIV-1 infection levels, and CCR5 ligands' concentrations in semen. SP+ and SP- displayed similar enhancing activity on PBMC infection by X4-tropic HIV-1. Addition/depletion of RANTES from seminal plasmas modulated their effect on PBMC infection by R5-tropic HIV-1. CONCLUSIONS: Semen from HIV-infected donors exhibit a significantly reduced enhancing potential on CD4+ T cell infection by R5-tropic HIV-1 when compared with semen from uninfected donors. Our data indicate that elevated seminal concentrations of RANTES in HIV+ men can influence the ability of semen to enhance infection.This is an open-access article distributed under the terms of the Creative Commons Attribution-Non Commercial-No Derivatives 4.0 License, where it is permissible to download and share the work provided it is properly cited. The work cannot be changed in any way or used commercially. http://creativecommons.org/licenses/by-nc-nd/4.0
AIDS

hal-01274885
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01274885
DOI : 10.1097/QAD.0000000000001048
PUBMED : 26854806

Éditeur(s) : HAL CCSD Portland Press
Résumé : International audience
CP12 is a small chloroplastic protein involved in the Calvin cycle that was shown to bind copper, a metal ion involved in modulation of its transition from reduced to oxidized state. In order to describe CP12 copper binding properties, copper-IMAC experiments and site-directed mutagenesis based on computational modeling, were coupled to top-down mass spectrometry (ESI-MS and MSMS). Copper-IMAC experiments allowed the primary characterization of mutation effects upon copper binding. Top-down MSMS experiments carried out under non-denaturing conditions on wild-type and mutant CP12/Cu2+ complexes then allowed fragment ions specifically liganding the copper ion to be determined. Comparison of MSMS datasets defined three regions involved in metal ion binding: residues D16 to D23, D38 to K50 and D70 to E76, with the two first regions containing selected residues for mutation. These data confirmed that copper ligands involved glutamic and aspartic residues in contrast to typical protein copper chelators. We propose that copper might play a role in regulation of CP12 biological activity.
ISSN: 0264-6021

hal-00479106
https://hal.archives-ouvertes.fr/hal-00479106
https://hal.archives-ouvertes.fr/hal-00479106/document
https://hal.archives-ouvertes.fr/hal-00479106/file/PEER_stage2_10.1042%252FBJ20082004.pdf
DOI : 10.1042/BJ20082004

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
The discerning behavior of living systems relies on accurate interactions selected from the lot of molecular collisions occurring in the cell. To ensure the reliability of interactions, binding partners are classically envisioned as finely preadapted molecules, selected on the basis of their affinity in one-step associations. But the counterselection of inappropriate interactions can in fact be much more efficiently obtained through difficult multi-step adjustment, whose final high energy state is locked by a fluctuation ratchet. The progressive addition of molecular bonds during stereo-adjustment can be modeled as a predominantly backward random walk whose first arrival is frozen by a micro-irreversible transition. A new criterion of ligand specificity is presented, that is based on the ratio rejection/incorporation. In addition to its role in the selectivity of interactions, this generic recipe can underlie other important biological phenomena such as the regular synthesis at low level of supramolecular complexes, monostable kinetic bimodality, substrate concentration thresholds or the preparation of rapidly depolymerizable structures with stored energy, like microtubules
ISSN: 0079-6107

hal-01274234
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01274234
DOI : 10.1016/j.pbiomolbio.2016.02.001
PUBMED : 26867859

Éditeur(s) : HAL CCSD Nature Publishing Group
Résumé : International audience
Activation of cell signaling by reactive chemicals and pollutants is an important issue for human health. It has been shown that lipophilic nitro-benzoxadiazole (NBD) compounds rapidly move across the plasma membrane and enhance Epidermal Growth Factor Receptor (EGFR) tyrosine phosphorylation in cancer cells. Unlike ligand-dependent activation, the mechanism of this induction relies on the generation of hydrogen peroxide, which is involved in the activation of the catalytic site of the receptor and the inactivation of protein tyrosine phosphatase PTP-1B. Production of H2O2 during redox transformation of NBD compounds is associated with the transition of a monomeric form of Cu/Zn superoxide dismutase 1 (SOD1) to stable dimers. The highly stable and functionally active SOD1 dimer, in the absence of adequate activities in downstream reactions, promotes the disproportionate production and accumulation of intracellular hydrogen peroxide shortly after exposure to NBD compounds. The intrinsic fluorescence of small compounds was used to demonstrate their binding to SOD1. Our data indicate that H2O2 and concomitantly generated electrophilic intermediates behave as independent entities, but all contribute to the biological reactivity of NBD compounds. This study opens a promising path to identify new biomarkers of oxidative/electrophilic stress in the progression of cancer and other diseases
ISSN: 2045-2322

hal-01279094
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01279094
DOI : 10.1038/srep21088
PUBMED : 26883293

Éditeur(s) : HAL CCSD Public Library of Science
Résumé : International audience
The Hedgehog (Hh) family of secreted proteins act as morphogens to control embryonic patterning and development in a variety of organ systems. Post-translational covalent attachment of cholesterol and palmitate to Hh proteins are critical for multimerization and long range signaling potency. However, the biological impact of lipid modifications on Hh ligand distribution and signal reception in humans remains unclear. In the present study, we report a unique case of autosomal recessive syndromic 46,XY Disorder of Sex Development (DSD) with testicular dysgenesis and chondrodysplasia resulting from a homozygous G287V missense mutation in the hedgehog acyl-transferase (HHAT) gene. This mutation occurred in the conserved membrane bound O-acyltransferase (MBOAT) domain and experimentally disrupted the ability of HHAT to palmitoylate Hh proteins such as DHH and SHH. Consistent with the patient phenotype, HHAT was found to be expressed in the somatic cells of both XX and XY gonads at the time of sex determination, and Hhat loss of function in mice recapitulates most of the testicular, skeletal, neuronal and growth defects observed in humans. In the developing testis, HHAT is not required for Sertoli cell commitment but plays a role in proper testis cord formation and the differentiation of fetal Leydig cells. Altogether, these results shed new light on the mechanisms of action of Hh proteins. Furthermore, they provide the first clinical evidence of the essential role played by lipid modification of Hh proteins in human testicular organogenesis and embryonic development.
ISSN: 1553-7390

hal-01063929
https://hal.archives-ouvertes.fr/hal-01063929
DOI : 10.1371/journal.pgen.1004340
PUBMED : 24784881

Éditeur(s) : HAL CCSD American Society for Biochemistry and Molecular Biology
Résumé : International audience
Neuroglobins, previously thought to be restricted to vertebrate neurons, were detected in the brain of a photosymbiotic acoel, Symsagittifera roscoffensis, and in neurosensory cells of the jellyfish Clytia hemisphaerica. For the neuroglobin of S. roscoffensis, a member of a lineage that originated either at the base of the bilateria or of the deuterostome clade, we report the ligand binding properties, crystal structure at 2.3 Å, and brain immunocytochemical pattern. We also describe in situ hybridizations of two neuroglobins specifically expressed in differentiating nematocytes (neurosensory cells) and in statocytes (ciliated mechanosensory cells) of C. hemisphaerica, a member of the early branching animal phylum cnidaria. In silico searches using these neuroglobins as queries revealed the presence of previously unidentified neuroglobin-like sequences in most metazoan lineages. Because neural systems are almost ubiquitous in metazoa, the constitutive expression of neuroglobin-like proteins strongly supports the notion of an intimate association of neuroglobins with the evolution of animal neural systems and hints at the preservation of a vitally important function. Neuroglobins were probably recruited in the first protoneurons in early metazoans from globin precursors. Neuroglobins were identified in choanoflagellates, sponges, and placozoans and were conserved during nervous system evolution. Because the origin of neuroglobins predates the other metazoan globins, it is likely that neuroglobin gene duplication followed by co-option and subfunctionalization led to the emergence of globin families in protostomes and deuterostomes (i.e. convergent evolution).
ISSN: 0021-9258

hal-01020584
http://hal.upmc.fr/hal-01020584
http://hal.upmc.fr/hal-01020584/document
http://hal.upmc.fr/hal-01020584/file/Rosko_jbc_revised.pdf
DOI : 10.1074/jbc.M112.407601

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
The Liver X Receptors (LXRs) α and β and the Peroxisome Proliferator-Activated Receptor α (PPARα) are transcription factors that belong to class II nuclear receptors. They drive the expression of genes involved in hepatic lipid homeostasis and therefore are important targets for the prevention and treatment of nonalcoholic fatty liver disease (NAFLD). LXRs and PPARα are regulated by endogenous ligands, oxysterols and fatty acid derived molecules, respectively. In the liver, pharmacological activation of LXRs leads to the over-expression of genes involved in de novo lipogenesis, while PPARα is critical for fatty acid catabolism in nutrient deprivation. Even if these two nuclear receptors seemed to play opposite parts, recent studies have highlighted that PPARα also influence the expression of genes involved in fatty acids synthesis. In this study, we used pharmacological approaches and genetically engineered mice to investigate the cross-talk between LXRs and PPARα in the regulation of genes responsible for lipogenesis. We first investigated the effect of T0901317 and fenofibrate, two synthetic agonists of LXRs and PPARα, respectively. As expected, T0901317 and fenofibrate induce expression of genes involved LXR-dependent and PPARα-dependent lipogenic responses. Considering such overlapping effect, we then tested whether LXR agonist may influence PPARα driven response and vice versa. We show that the lack of PPARα does not influence the effects of T0901317 on lipogenic genes expression. However, PPARα deficiency prevents the up-regulation of genes involved in ω-hydroxylation that are induced by the LXR agonist. In addition, over-expression of lipogenic genes in response to fenofibrate is decreased in LXR knockout mice as well as the expression of PPARα target genes involved in fatty acid oxidation. Altogether, our work provides in vivo evidence for a central interconnection between nuclear receptors that drive hepatic lipid metabolism in response to oxysterol and fatty acids.
ISSN: 0300-9084

inserm-00872582
http://www.hal.inserm.fr/inserm-00872582
DOI : 10.1016/j.biochi.2012.09.028
PUBMED : 23063693

Éditeur(s) : HAL CCSD Frontiers
Résumé : International audience
Important set of studies have demonstrated the endocrine disrupting activity of Bisphenol A (BPA). The present work aimed at defining estrogenic-like activity of several BPA structural analogs, including BPS, BPF, BPAF, and BPAP, on 4- or 7-day post-fertilization (dpf) zebrafish larva as an in vivo model. We measured the induction level of the estrogen-sensitive marker cyp19a1b gene (Aromatase B), expressed in the brain, using three different in situ/in vivo strategies: (1) Quantification of cyp19a1b transcripts using RT-qPCR in wild type 7-dpf larva brains exposed to bisphenols; (2) Detection and distribution of cyp19a1b transcripts using in situ hybridization on 7-dpf brain sections (hypothalamus); and (3) Quantification of the cyp19a1b promoter activity in live cyp19a1b-GFP transgenic zebrafish (EASZY assay) at 4-dpf larval stage. These three different experimental approaches demonstrated that BPS, BPF, or BPAF exposure, similarly to BPA, significantly activates the expression of the estrogenic marker in the brain of developing zebrafish. In vitro experiments using both reporter gene assay in a glial cell context and competitive ligand binding assays strongly suggested that up-regulation of cyp19a1b is largely mediated by the zebrafish estrogen nuclear receptor alpha (zfERα). Importantly, and in contrast to other tested bisphenol A analogs, the bisphenol AP (BPAP) did not show estrogenic activity in our model
ISSN: 1662-4548

hal-01299976
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01299976
PUBMED : 27047331
DOI : 10.3389/fnins.2016.00112
PUBMEDCENTRAL : PMC4805609

Éditeur(s) : HAL CCSD
Résumé : contient des versions PDF d'articles scientifiques (versions éditeur)
Spectrins are actin binding proteins that constitute the main components of membrane skeletons, the protein network located under the plasma membrane. Erythroid or non erythroid (in this case, they are called fodrins), spectrins play an important structural function in the membrane as demonstrated in the red blood cell. Spectrins are composed of two long α and β chains, associated in tetramers (αβ)2, forming the long flexible filaments of the network. Both chains are composed of repeating units which are folded in a triple helical arrangement (A, B and C helices), except for segment α10 which corresponds to a Src Homology 3 domain (SH3). The head-to-head interaction between dimers implies the N-terminal end of α chain and the C-terminal end of β chain. A model was proposed for the tetramerization site of spectrin : the two A and B helices of the last repeat of α chain interact with the C helix of the β chain in order to constitute a triple-helical structure similar to the repeats along the remainder of spectrin molecule. This model was first inferred from the relationship between the severity of the spectrin self-association defect and the location of the underlying mutations found in hereditary elliptocytosis (HE). Using recombinant peptides we defined the region, on both chains, which was sufficient and necessary to generate a fully functional tetramerization site. Producing peptides carrying a βHE mutation, we reproduced the tetramerization defect as observed in red blood cells of patients.
Finally, the presence of an SH3 domain probably gives to spectrins new functions which are different from the stability of the membrane. After contesting the existence of interaction between the fodrin SH3 domain and the α subunit of epithelial sodium channel sensitive to amiloride, we searched for partners of this domain, by screening a rat kidney library with the yeast two-hybrid system. We identified severals proteins implicated in the actin polymerization asoociated with motility or cell differenciation, suggesting that SH3 domain could play a part in this process.
Les spectrines, protéines liant l'actine, sont des constituants majeurs du squelette membranaire, réseau multiprotéique localisé sous la membrane plasmique. Érythroïdes ou non érythroïdes (dans ce cas, on les appelle fodrines), les spectrines ont un rôle structural important dans la membrane comme cela a déjà été démontré, pour la spectrine, dans le globule rouge. Elles sont constituées de deux longues chaînes a et ß associées côte à côte en tétramères (aß)2 qui forment les longs filaments du réseau. Chacune des chaînes est composée par la répétition de segments homologues, 22 pour a et 17 pour ß. Ces segments sont constitués de trois hélices a (hélices A, B et C) repliées sur elles-mêmes. Cette succession de structures trihélicoïdales est parfois interrompue par un domaine particulier comme le domaine SH3 (Src Homology 3) présent au milieu de la chaîne a. Les tétramères (aß)2 de spectrine constituent les filaments du squelette membranaire. L'interaction tête-à-tête de deux dimères aß implique les extrémités NH2 de la chaîne a et COOH de la chaîne ß. D'une part la sévérité du défaut d'auto-association et la localisation des mutations associées à celui-ci et d'autre part, les séquences en acides aminés des extrémités impliquées dans le site d'auto-association ont permis de proposer un modèle : la première hélice C isolée de la chaîne a pourrait s'associer aux deux dernières hélices A et B de la chaîne ß pour reconstituer une unité conformationnelle trihélicoïdale semblable à celles observées le long de la molécule. Un défaut dans la formation du tétramère est le support moléculaire le plus fréquemment observé dans les elliptocytoses héréditaires (HE). À l'aide de peptides recombinants, nous avons défini, sur les deux chaînes, les régions nécessaires et suffisantes possédant les caractéristiques pleinement fonctionnelles du site de tétramérisation. Nous avons ensuite, par mutagenèse dirigée, reproduit le lien entre la présence de mutations HE localisées dans les hélices A ou B et le défaut d'auto-association observé dans les globules rouges de patients HE. La présence d'un domaine SH3 localisé au milieu de la chaîne a confère probablement aux spectrine des fonctions autres que le maintien et la stabilité de la membrane. Les SH3, petits domaines protéiques, participent aux interactions protéine/protéine. Le seul partenaire connu du domaine SH3 de la fodrine était la sous-unité a du canal sodium sensible à l'amiloride (ENaC) mais la fonction de ce complexe n'était pas encore caractérisée. À l'aide de différentes méthodes, nous avons remis en cause l'interaction directe entre ENaC et le SH3 de la fodrine. La fonction de ce domaine SH3 étant liée à la nature de son ligand, nous avons donc recherché les partenaires putatifs du domaine SH3 de la fodrine par la technique du double-hybride. 29 partenaires ont été identifiés, regroupés en 19 familles et 10 clones isolés. La spécificité des interactions a été étudiée à la fois par double-hybride, à l'aide de mutants du domaine SH3 de la fodrine produits par mutagenèse dirigée. Les interactions vis-à-vis d'autres domaines SH3 (spectrine ou une protéine de levure non relatée Scd2) ont été également analysées. Enfin, la spécificité de certains partenaires a été confirmée par des études d'interactions in vitro. Deux des protéines les plus spécifiques du domaine SH3 de la fodrine sont des protéines tyrosine-phosphatase PTP. La première est l'isoforme A de PTP de faible poids moléculaire (LMPTPA) mais l'interaction n'a pas été confirmée in vitro. La deuxième, TD14, est une nouvelle PTP dont la seule fonction connue est d'inhiber la formation des foyers tumoraux des cellules surexprimant l'oncogène Ha-ras. Ces PTP pourraient soit déphosphoryler la fodrine, soit être recrutées sous la membrane pour déphosphoryler une autre cible. Nous avons également identifié trois partenaires (N-WASP, Evl et une protéine de la famille des formines) suggérant que les domaines SH3 des spectrines pourraient être impliqués dans les processus de polymérisation de l'actine liés à la mobilité ou la différenciation cellulaire. Mot clés : spectrine, fodrine, tétramérisation, SH3, double-hybride, polymérisation de l'actine
https://tel.archives-ouvertes.fr/tel-00284819

Droits : info:eu-repo/semantics/OpenAccess
tel-00284819
https://tel.archives-ouvertes.fr/tel-00284819
https://tel.archives-ouvertes.fr/tel-00284819/document
https://tel.archives-ouvertes.fr/tel-00284819/file/Nicolas_1999_-_These.pdf

Éditeur(s) : HAL CCSD
Résumé : International audience
Millimeter waves (MMW) will be increasingly used for future wireless telecommunications. Previous studies on skin keratinocytes showed that MMW could impact the mRNA expression of Transient Receptor Potential cation channel subfamily Vanilloid, member 2 (TRPV2). Here, we investigated the effect of MMW exposure on this marker, as well as on other membrane receptors such as Transient Receptor Potential cation channel subfamily Vanilloid, member 1 (TRPV1) and purinergic receptor P2X, ligand-gated ion channel, 3 (P2 × 3). We exposed the Neuroscreen-1 cell line (a PC12 subclone), in order to evaluate if acute MMW exposures could impact expression of these membrane receptors at the protein level. Proteotoxic stress-related chaperone protein Heat Shock Protein 70 (HSP70) expression level was also assessed. We used an original high-content screening approach, based on fluorescence microscopy, to allow cell-by-cell analysis and to detect any cell sub-population responding to exposure. Immunocytochemistry was done after 24 h MMW exposure of cells at 60.4 GHz, with an incident power density of 10 mW/cm2. Our results showed no impact of MMW exposure on protein expressions of HSP70, TRPV1, TRPV2, and P2 × 3. Moreover, no specific cell sub-populations were found to express one of the studied markers at a different level, compared to the rest of the cell populations. However, a slight insignificant increase in HSP70 expression and an increase in protein expression variability within cell population were observed in exposed cells, but controls showed that this was related to thermal effect. Bioelectromagnetics. 37:444–454, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.
Bioelectromagnetics

hal-01371963
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01371963
DOI : 10.1002/bem.21995
PUBMED : 27483046

Éditeur(s) : HAL CCSD
Résumé : Mon document "Titres et travaux" contient 5 articles au format éditeur.
Mes premiers travaux de recherche, me permettant de soutenir une thèse de doctorat en décembre 1999, ont été effectués au sein du laboratoire du professeur Bernard GRANDCHAMP (unité 409 de l'INSERM) sous la direction du docteur Marie-Christine LECOMTE. Mon travail s'est intégré dans l'étude fonctionnelle et structurale de certains domaines des spectrines érythroïdes et non érythroïdes, principalement le site de tétramérisation et le domaine SH3. Concernant la tétramérisation de la spectrine érythroïde j'ai identifié les régions minimales nécessaires et suffisantes pour former le tétramère puis j'ai mis en évidence une relation entre certaines mutations de la spectrine associées à des anomalies membranaires du globule rouge et la sévérité du défaut de formation du tétramère. J'ai ensuite identifié plusieurs ligands du domaine SH3 de la spectrine non érythroïde. L'un d'entre eux m'a permis de définir une régulation de la protéolyse ciblée de cette spectrine faisant intervenir un mécanisme de phosphorylation/déphosphorylation. Ma formation initiale fut donc celle d'un biochimiste que j'ai étendue peu à peu vers la biologie moléculaire et la biologie cellulaire.
Puis, souhaitant acquérir les connaissances nécessaires au développement de modèles murins, j'ai décidé d'effectuer mon stage post-doctoral à l'Institut COCHIN (unité 567 de l'INSERM) dans une équipe co-dirigée par le docteur Sophie VAULONT et le professeur Axel KAHN. Je me suis ainsi familiarisé avec la souris comme « outil » de recherche (gestion d'un élevage, établissement de nouvelles lignées, génotypage, phénotypage). Je travaillais initialement sur le métabolisme du glucose mais j'ai complètement réorienté ma thématique de recherche au bout d'un an pour finalement identifier et caractériser une hormone, l'hepcidine, contrôlant l'homéostasie du fer.
J'ai été nommé chargé de recherche de grade 2 dans l'unité 409 de l'INSERM en octobre 2002 pour y développer une souche de souris exprimant une spectrine non érythroïde résistante à des mécanismes de protéolyse ciblée. Il s'agit donc toujours d'appréhender certaines fonctions des spectrines mais dans un contexte in vivo. En janvier 2005 j'ai intégré l'unité 665 de l'INSERM dirigée par Yves COLIN afin de poursuivre ce projet. Aujourd'hui, je suis chargé de recherche de grade 1.
Le chapitre scientifique de ce mémoire est divisé en deux parties distinctes : la première traite de mon travail sur la spectrine tandis que la seconde porte sur l'hepcidine. Chaque partie est composée d'une introduction au sujet suivie d'un résumé des travaux que j'ai effectués et/ou dirigés.
https://tel.archives-ouvertes.fr/tel-00285035

Droits : info:eu-repo/semantics/OpenAccess
tel-00285035
https://tel.archives-ouvertes.fr/tel-00285035
https://tel.archives-ouvertes.fr/tel-00285035/document
https://tel.archives-ouvertes.fr/tel-00285035/file/HDR_Titre_et_travaux_-_Gael_Nicolas.pdf

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
Apoptosis is a fundamental process that contributes to tissue homeostasis, immune responses, and development. The receptor CD95, also called Fas, is a member of the tumor necrosis factor receptor (TNF-R) superfamily. Its cognate ligand, CD95L, is implicated in immune homeostasis and immune surveillance, and various lineages of malignant cells exhibit loss-of-function mutations in this pathway; therefore, CD95 was initially classified as a tumor suppressor gene. However, more recent data indicate that in different pathophysiological contexts, this receptor can transmit non-apoptotic signals, promote inflammation, and contribute to carcinogenesis. A comparison with the initial molecular events of the TNF-R signaling pathway leading to non-apoptotic, apoptotic, and necrotic pathways reveals that CD95 is probably using different molecular mechanisms to transmit its non-apoptotic signals (NF-κB, MAPK, and PI3K). As discussed in this review, the molecular process by which the receptor switches from an apoptotic function to an inflammatory role is unknown. More importantly, the biological functions of these signals remain elusive.
ISSN: 0006-3002

hal-01061290
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01061290
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01061290/document
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01061290/file/BBAreviewsoncancer-legembre-final.pdf
DOI : 10.1016/j.bbcan.2014.04.007

Éditeur(s) : HAL CCSD Springer Verlag
Résumé : International audience
Bone Morphogenetic Protein 9 (BMP9) has been recently found to be the physiological ligand for the activin receptor-like kinase 1 (ALK1), and to be a major circulating vascular quiescence factor. Moreover, a soluble chimeric ALK1 protein (ALK1-Fc) has recently been developed and showed powerful anti-tumor growth and anti-angiogenic effects. However, not much is known concerning BMP9. This prompted us to investigate the human endogenous sources of this cytokine and to further characterize its circulating form(s) and its function. Analysis of BMP9 expression reveals that BMP9 is produced by hepatocytes and intrahepatic biliary epithelial cells. Gel filtration analysis combined with ELISA and biological assays demonstrate that BMP9 circulates in plasma (1) as an unprocessed inactive form that can be further activated by furin a serine endoprotease, and (2) as a mature and fully active form (composed of the mature form associated with its prodomain). Analysis of BMP9 circulating levels during mouse development demonstrates that BMP9 peaks during the first 3 weeks after birth and then decreases to 2 ng/mL in adulthood. We also show that circulating BMP9 physiologically induces a constitutive Smad1/5/8 phosphorylation in endothelial cells. Taken together, our results argue for the role of BMP9 as a hepatocyte-derived factor, circulating in inactive (40%) and active (60%) forms, the latter constantly activating endothelial cells to maintain them in a resting state.
ISSN: 1420-682X

hal-00681969
https://hal.archives-ouvertes.fr/hal-00681969
DOI : 10.1007/s00018-011-0751-1
PUBMED : 21710321

Éditeur(s) : HAL CCSD American Society for Biochemistry and Molecular Biology
Résumé : International audience
Polycyclic aromatic hydrocarbons (PAHs) such as benzo(a)pyrene (B(a)P) are widely distributed environmental contaminants, known as potent ligands of the aryl hydrocarbon receptor (AhR). These chemicals trigger an early and transient increase of intracellular calcium concentration ([Ca(2+)](i)), required for AhR-related effects of PAHs. The mechanisms involved in this calcium mobilization were investigated in the present study. We demonstrated that B(a)P-mediated [Ca(2+)](i) induction was prevented in endothelial HMEC-1 cells by counteracting β2-adrenoreceptor (β2ADR) activity using pharmacological antagonists, anti-β2ADR antibodies, or siRNA-mediated knockdown of β2ADR expression; by contrast, it was strongly potentiated by β2ADR overexpression in human kidney HEK293 cells. B(a)P was shown, moreover, to directly bind to β2ADR, as assessed by in vitro binding assays and molecular modeling. Pharmacological inhibition and/or siRNA-mediated silencing of various signaling actors acting downstream of β2ADR in a sequential manner, such as G protein, adenylyl cyclase, Epac-1 protein, and inositol 1,4,5-trisphosphate (IP(3))/IP(3) receptor, were next demonstrated to prevent B(a)P-induced calcium signal. Inhibition or knockdown of these signaling elements, as well as the use of chemical β-blockers, were finally shown to counteract B(a)P-mediated induction of cytochrome P-450 1B1, a prototypical AhR target gene. Taken together, our results show that B(a)P binds directly to β2ADR and consequently utilizes β2ADR machinery to mobilize [Ca(2+)](i), through activation of a G protein/adenylyl cyclase/cAMP/Epac-1/IP(3) pathway. This β2ADR-dependent signaling pathway activated by PAHs may likely be crucial for PAH-mediated up-regulation of AhR target genes, thus suggesting a contribution of β2ADR to the health-threatening effects of these environmental pollutants.
ISSN: 0021-9258

hal-00682497
https://hal.archives-ouvertes.fr/hal-00682497
DOI : 10.1074/jbc.M111.319970
PUBMED : 22167199

Éditeur(s) : HAL CCSD Elsevier
Résumé : International audience
CD95 ligand (CD95L) is expressed by immune cells and triggers apoptotic death. Metalloprotease-cleaved CD95L (cl-CD95L) is released into the bloodstream but does not trigger apoptotic signaling. Hence, the pathophysiological role of cl-CD95L remains unclear. We observed that skin-derived endothelial cells from systemic lupus erythematosus (SLE) patients expressed CD95L and that after cleavage, cl-CD95L promoted T helper 17 (Th17) lymphocyte transmigration across the endothelial barrier at the expense of T regulatory cells. T cell migration relied on a direct interaction between the CD95 domain called calcium-inducing domain (CID) and the Src homology 3 domain of phospholipase C gamma 1. Th17 cells stimulated with cl-CD95L produced sphingosine-1-phosphate (S1P), which promoted endothelial transmigration by activating the S1P receptor 3. We generated a cell-penetrating CID peptide that prevented Th17 cell transmigration and alleviated clinical symptoms in lupus mice. Therefore, neutralizing the CD95 non-apoptotic signaling pathway could be an attractive therapeutic approach for SLE treatment.
ISSN: 1074-7613

hal-01359568
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01359568
DOI : 10.1016/j.immuni.2016.06.028
PUBMED : 27438772

Éditeur(s) : HAL CCSD Impact journals
Résumé : International audience
TNF-Related Apoptosis-Inducing Ligand (TRAIL) is a well-known apoptosis inducer, which activates the extrinsic death pathway. TRAIL is pro-apoptotic on colon cancer cells, while not cytotoxic towards normal healthy cells. However, its clinical use is limited by cell resistance to cell death which occurs in approximately 50% of cancer cells. Short Chain Fatty Acids (SCFA) are also known to specifically induce apoptosis of cancer cells. In accordance, we have shown that food grade dairy propionibacteria induce intrinsic apoptosis of colon cancer cells, via the production and release of SCFA (propionate and acetate) acting on mitochondria. Here, we investigated possible synergistic effect between Propionibacterium freudenreichii and TRAIL. Indeed, we hypothesized that acting on both extrinsic and intrinsic death pathways may exert a synergistic pro-apoptotic effect. Whole transcriptomic analysis demonstrated that propionibacterial supernatant or propionibacterial metabolites (propionate and acetate), in combination with TRAIL, increased pro-apoptotic gene expression (TRAIL-R2/DR5) and decreased anti-apoptotic gene expression (FLIP, XIAP) in HT29 human colon cancer cells. The revealed synergistic pro-apoptotic effect, depending on both death receptors (TRAIL-R1/DR4, TRAIL-R2/DR5) and caspases (caspase-8, -9 and -3) activation, was lethal on cancer cells but not on normal human intestinal epithelial cells (HIEC), and was inhibited by Bcl-2 expression. Finally, milk fermented by P. freudenreichii induced HT29 cells apoptosis and enhanced TRAIL cytotoxic activity, as did P. freudenreichii DMEM culture supernatants or its SCFA metabolites. These results open new perspectives for food grade P. freudenreichii-containing products in order to potentiate TRAIL-based cancer therapy in colorectal cancer
ISSN: 1949-2553

hal-01259215
https://hal-univ-rennes1.archives-ouvertes.fr/hal-01259215
DOI : 10.18632/oncotarget.6881
PUBMED : 26771233