Altered Ca2+ Homeostasis in Red Blood Cells of Polycythemia Vera Patients Following Disturbed Organelle Sorting during Terminal Erythropoiesis

The authors thank Thierry Peyrard, Dominique Gien, Sirandou Tounkara, and Eliane Véra at Centre National de Référence pour les Groupes Sanguins for the management of blood samples. The authors thank Sandrine Genetet and Isabelle Mouro-Chanteloup at the Inserm UMR_S1134 unit for their assistance in experiments. The authors also thank Michaël Dussiot at the Institute Imagine for his assistance in imaging flow cytometry. We thank Johanna Bruce and Virginie Salnot at 3P5 Proteomics Platform for sample preparation and analysis, and François Guillonneau and Patrick Mayeux for their management and strategies. Funding: The work was supported by Institut National de la Santé et de la Recherche Médicale (Inserm); Institut National de la Transfusion Sanguine (INTS); the University of Paris; and grants from Laboratory of Excellence (Labex) GR-Ex, reference No. ANR-11-LABX-0051. The Labex GR- Ex is funded by the IdEx program “Investissements d’avenir” of the French National Research Agency, reference No. ANR-11-IDEX-0005-02 and ANR-18-IDEX-0001. R.B., M.G.R., and D.M.A. were funded by the European Union’s Horizon 2020 Research and Innovation Program under grant agreement No. 675115-RELEVANCE-H2020-MSCA-ITN-2015. R.B. also received financial support from Société Française d’Hématologie (SFH) and Club du Globule Rouge et du Fer (CGRF). R.B. is currently funded by the Innovate UK Research and Innovation Knowledge Transfer Partnership (KTP) between University of Aberdeen and Vertebrate Antibodies Ltd. (Partnership No. KTP12327). T.D. was supported by PhD grants from Université Paris Saclay MESR (Ministère Enseignement Supérieur et de la Recherche) and then FRM (Fondation recherche médicale). The Orbitrap Fusion mass spectrometer was acquired with funds from Fonds Europeen de Developpement Regional (FEDER) through the Operational Program for Competitiveness Factors and Employment 2007-2013 and from the Canceropole Ile de France.Peer reviewedPublisher PD

Optimisation de la cartographie peptidique MALDI-TOF par addition d'éther couronne

Diplôme : Licence Professionnell

Optimisation de la cartographie peptidique par addition d'éther couronne 18-C-6

National audienc

Pharmacological profile of engineered 5-HT4 receptors and identification of 5-HT4 receptor-biased ligands

International audienceG protein-coupled receptors (GPCRs) can activate simultaneously multiple signaling pathways upon agonist binding. The combined use of engineered GPCRs, such as the Receptors Activated Solely by Synthetic Ligands (RASSLs), and of biased ligands that activate only one pathway at a time might help deciphering the physiological role of each G protein signaling. In order to find serotonin type 4 receptor (5-HT4R) biased ligands, we analyzed the ability of several compounds to activate the Gs and Gq/11 pathways in COS-7 cells that transiently express wild type 5-HT4R, the 5-HT4R-D 100 A mutant (known also as 5-HT4-RASSL, or Rs1) or the 5-HT4R-T 104 A mutant, which modifies agonist-induced 5-HT4R activation. This analysis allowed completing the pharmacological profile of the two mutant 5-HT4Rs, but we did not find any biased ligand for the mutant receptors. Conversely, we identified the first biased agonists for wild type 5-HT4R. Indeed, RS 67333 and prucalopride acted as partial agonists to induce cAMP accumulation, but as antagonists on inositol phosphate production. Moreover, they showed very different antagonist potencies that could be exploited to study the activation of the Gs pathway, with or without concomitant block of Gq/11 signaling

New Interactors of the Truncated EBNA-LP Protein Identified by Mass Spectrometry in P3HR1 Burkitt’s Lymphoma Cells

The Epstein-Barr virus nuclear antigen leader protein (EBNA-LP) acts as a co-activator of EBNA-2, a transcriptional activator essential for Epstein-Barr virus (EBV)-induced B-cell transformation. Burkitt’s lymphoma (BL) cells harboring a mutant EBV strain that lacks both the EBNA-2 gene and 3′ exons of EBNA-LP express Y1Y2-truncated isoforms of EBNA-LP (tEBNA-LP) and better resist apoptosis than if infected with the wild-type virus. In such BL cells, tEBNA-LP interacts with the protein phosphatase 2A (PP2A) catalytic subunit (PP2A C), and this interaction likely plays a role in resistance to apoptosis. Here, 28 cellular and four viral proteins have been identified by mass spectrometry as further possible interactors of tEBNA-LP. Three interactions were confirmed by immunoprecipitation and Western blotting, namely with the A structural subunit of PP2A (PP2A A), the structure-specific recognition protein 1 (SSRP1, a component of the facilitate chromatin transcription (FACT) complex), and a new form of the transcription factor EC (TFEC). Thus, tEBNA-LP appears to be involved not only in cell resistance to apoptosis through its interaction with two PP2A subunits, but also in other processes where its ability to co-activate transcriptional regulators could be important

ZNF555 protein binds to transcriptional activator site of 4qA allele and ANT1: potential implication in Facioscapulohumeral dystrophy

International audienceFacioscapulohumeral dystrophy (FSHD) is an epi/genetic satellite disease associated with at least two satellite sequences in 4q35: (i) D4Z4 macrosatellite and (ii) β-satellite repeats (BSR), a prevalent part of the 4qA allele. Most of the recent FSHD studies have been focused on a DUX4 transcript inside D4Z4 and its tandem contraction in FSHD patients. However, the D4Z4-contraction alone is not pathological, which would also require the 4qA allele. Since little is known about BSR, we investigated the 4qA BSR functional role in the transcriptional control of the FSHD region 4q35. We have shown that an individual BSR possesses enhancer activity leading to activation of the Adenine Nucleotide Translocator 1 gene (ANT1), a major FSHD candidate gene. We have identified ZNF555, a previously uncharacterized protein, as a putative transcriptional factor highly expressed in human primary myoblasts that interacts with the BSR enhancer site and impacts the ANT1 promoter activity in FSHD myoblasts. The discovery of the functional role of the 4qA allele and ZNF555 in the transcriptional control of ANT1 advances our understanding of FSHD pathogenesis and provides potential therapeutic targets

Les risques conchylicoles en Baie de Quiberon. Première partie : le risque de mortalité virale du naissain d’huître creuse Crassostra gigas. Rapport final du projet Risco 2010-2013

This study (“Risco”), implicating both industry, socio-economic experts and biologists, was funded by the Regional Council of Brittany, for 3 years (2010-2013), to investigate upon the origin of oysters (Crassostrea gigas) mortalities in the bay of Quiberon (South Brittany, France). Specific mortalities of adult oysters were recorded in this bay in 2006, whereas mortalities of one year old spat were observed since in 2008 in this bay as at a national scale. The experimental protocol in 2010, including a monthly survey of oyster batches at 15 experimental stations, allowed to assess the risk for one-year oysters, due to viral disease. A clear spatial distribution of this risk was evidenced, with a western area spared from contamination and mortality, and a central and north-east sector strongly affected. The virus analysis clearly demonstrate the responsibility of the OsHV-1 virus in the mortalities. The oysters on the bottom with a lower growth appear less sensitive to the viral disease than the surelevated ones. In order to interpret this spatial distribution, an epidemiological model has been tested : it is based on emission of virus from the stock of spat in surrounding farming areas, dispersion by the hydrodynamic conditions, and inactivation of the virus by solar radiation. The resulting simulations suggest that the contamination would be endogenous to the bay (from contaminated stocks of spat seeded nearby). A decrease of densities for rearing spat, as well as the seeding of non contaminated spat may be recommended.L’étude « Risco », labellisée Pôle Mer et financée par la Région Bretagne, mobilise à la fois des socio-économistes, des biologistes et des professionnels. Elle vise à comprendre et gérer les facteurs de mortalités massives d’huîtres creuses (Crassostrea gigas) enregistrées par les concessionnaires de baie de Quiberon (France, 56), à partir de 2006 sur les huîtres adultes et 2008 sur le naissain. Le protocole engagé en 2010, avec un volet expérimental basé sur le suivi mensuel de lots d’huîtres en 15 stations et des analyses pathologiques, a permis d’éclairer notamment le risque épizootique sur le naissain. Une spatialisation très marquée de ce risque a été mise en évidence, avec une zone à l’ouest relativement épargnée et une zone au centre et à l’est très affectée. Les analyses virales mettent clairement en évidence la responsabilité du virus OsHV-1 dans ces mortalités. Les huîtres élevées au sol, moins poussantes, seraient aussi moins sensibles à la mortalité virale que les huîtres élevées en surélévation. L’existence d’une zone quasi-indemne de contamination et de mortalité à cette échelle est inédite parmi les secteurs ostréicoles français, depuis 2008. Pour interpréter cette distribution spatiale de la contamination et des mortalités, un modèle épidémiologique a été testé : il s’appuie sur une émission de virus à partir des stocks de naissain estimés en 2010 (estran et eau profonde), une dispersion par les courants, et une inactivation du virus en fonction du rayonnement solaire. Avec le taux d’abattement viral retenu, les simulations suggèrent que la contamination serait majoritairement endogène à la baie (à partir de semis de naissain en place, contaminés). Les recommandations qui en découlent sont notamment d’introduire en baie du naissain non contaminé et de diminuer les densités de naissain

Absolute proteome quantification of highly purified populations of circulating reticulocytes and mature erythrocytes

International audienc

Molecular Architecture and Function of the SEA Complex, a Modulator of the TORC1 Pathway*

The TORC1 signaling pathway plays a major role in the control of cell growth and response to stress. Here we demonstrate that the SEA complex physically interacts with TORC1 and is an important regulator of its activity. During nitrogen starvation, deletions of SEA complex components lead to Tor1 kinase delocalization, defects in autophagy, and vacuolar fragmentation. TORC1 inactivation, via nitrogen deprivation or rapamycin treatment, changes cellular levels of SEA complex members. We used affinity purification and chemical cross-linking to generate the data for an integrative structure modeling approach, which produced a well-defined molecular architecture of the SEA complex and showed that the SEA complex comprises two regions that are structurally and functionally distinct. The SEA complex emerges as a platform that can coordinate both structural and enzymatic activities necessary for the effective functioning of the TORC1 pathway

Use of a serious game to strengthen medication adherence in euthymic patients with bipolar disorder following a psychoeducational programme: A randomized controlled trial

International audienceAlthough psychoeducation programmes are the gold-standard intervention in bipolar disorder (BD), more innovative tools are needed to broaden and consolidate their effects, especially on treatment adherence. Serious games could be an option. Methods: We carried out a two-arm open randomized controlled trial to compare the add-on use of the serious game BIPOLIFE® for one month (n = 20) vs. treatment as usual (TAU; n = 21) following the completion of a psychoeducation programme in euthymic adults with BD. The primary outcome was the percentage of adherent patients (i.e., patients with a Medication Adherence Rating Scale, MARS, total score >7) at 4 months after the end of the psychoeducation programme. We also measured the changes in therapeutic adherence and beliefs on pharmacological treatments (Drug Attitude Inventory, DAI) between study inclusion and the 1-month (end of BIPOLIFE® use) and 4-month visits, healthcare use during the study period, and BIPOLIFE® acceptability.Results: The percentage of adherent patients was lower in the BIPOLIFE® group than in the TAU group at inclusion (p = 0.02). Conversely, the absolute variation of the MARS and DAI scores was higher in the BIPOLIFE® than in the TAU group at the 1-month visit (p = 0.03 and p = 0.002, respectively) but not at the 4-month visit (p = 0.22 and p = 0.07, respectively).Limitations: Small sample size, and low frequency of connexion to BIPOLIFE® declared by the patients.Conclusion: BIPOLIFE® may help patients with BD to increase their confidence in medications, if used regularly