Orthoparamyxovirinae C Proteins Have a Common Origin and a Common Structural Organization

The protein C is a small viral protein encoded in an overlapping frame of the P gene in the subfamily Orthoparamyxovirinae. This protein, expressed by alternative translation initiation, is a virulence factor that regulates viral transcription, replication, and production of defective interfering RNA, interferes with the host-cell innate immunity systems and supports the assembly of viral particles and budding. We expressed and purified full-length and an N-terminally truncated C protein from Tupaia paramyxovirus (TupV) C protein (genus Narmovirus). We solved the crystal structure of the C-terminal part of TupV C protein at a resolution of 2.4 Å and found that it is structurally similar to Sendai virus C protein, suggesting that despite undetectable sequence conservation, these proteins are homologous. We characterized both truncated and full-length proteins by SEC-MALLS and SEC-SAXS and described their solution structures by ensemble models. We established a mini-replicon assay for the related Nipah virus (NiV) and showed that TupV C inhibited the expression of NiV minigenome in a concentration-dependent manner as efficiently as the NiV C protein. A previous study found that the Orthoparamyxovirinae C proteins form two clusters without detectable sequence similarity, raising the question of whether they were homologous or instead had originated independently. Since TupV C and SeV C are representatives of these two clusters, our discovery that they have a similar structure indicates that all Orthoparamyxovirine C proteins are homologous. Our results also imply that, strikingly, a STAT1-binding site is encoded by exactly the same RNA region of the P/C gene across Paramyxovirinae, but in different reading frames (P or C), depending on which cluster they belong to.French Agence Nationale de la RechercheFond de la Recherche Médicale (FRM)Grenoble Instruct-ERIC centerFRISBIUniversity Grenoble Alpes graduate school (Ecoles Universitaires de Recherche)Peer Reviewe

Gene expression profiling of human adrenocortical tumors using complementary deoxyribonucleic Acid microarrays identifies several candidate genes as markers of malignancy.

International audienceThe aim of this study was to identify predictor sets of genes whose over- or underexpression in human sporadic adrenocortical tumors would help to identify malignant vs. benign tumors and to predict postsurgical metastatic recurrence. For this, we analyzed the expression of 230 candidate genes using cDNA microarrays in a series of 57 well-characterized human sporadic adrenocortical tumors (33 adenomas and 24 carcinomas). We identified two clusters of genes (the IGF-II cluster containing eight genes, including IGF-II, and the steroidogenesis cluster containing six genes encoding steroidogenic enzymes plus eight other genes) whose combined levels of expression appeared to be good predictors of malignancy. This predictive value was as strong as that of the pathological score of Weiss. The analysis of the population of carcinomas (13 tumors) for genes whose expression would be strongly different between recurring and nonrecurring tumors allowed identification of 14 genes meeting these criteria. Among these genes, there are probably new markers of tumor evolution that will deserve additional validation on a larger scale. Taken together, these results show that the parallel analysis of the expression levels of a selected group of genes on microgram quantities of tumor RNA (a quantity that can be obtained from fine needle aspirations) appears as a complementary method to histopathology for the diagnosis and prognosis of evolution of adrenocortical carcinomas

Single and combined impacts of irradiation and surgery on lymphatic vasculature and fibrosis associated to secondary lymphedema

peer reviewedLymphedema (LD) refers to a condition of lymphatic dysfunction associated with excessive fluid accumulation, fibroadipose tissue deposition and swelling. In industrialized countries, LD development mainly results from a local disruption of the lymphatic network by an infection or cancer-related surgery (secondary LD). In the absence of efficient therapy, animal models are needed to decipher the cellular and molecular mechanisms underlying LD and test putative drugs. In this study, we optimized and characterized a murine model of LD that combines an irradiation of the mice hind limb and a radical surgery (lymph node resection associated to lymphatic vessel ligation). We investigated the respective roles of irradiation and surgery in LD formation by comparing their impacts, alone or in combination (with different intervention sequences), on eight different features of the pathology: swelling (paw thickness), indocyanine green (ICG) clearance, lymphatic vasculature remodeling, epidermal and dermal thickening, adipocyte accumulation, inflammatory cell infiltration and collagen deposition. This study supports the importance of radiation prior to surgery to experimentally induce a rapid, severe and sustained tissue remodeling harboring the different hallmarks of LD. We provide the first experimental evidence for an excessive deposition of periostin (POSTN) and tenascin-C (TNC) in LD. Through a computerized method of digital image quantification, we established the spatial map of lymphatic expansion, as well as collagen, POSTN and TNC deposition in papillary and reticular dermis of lymphedematous skins. This mouse model is available to study the patho-physiology of LD and test potential therapeutic targets

An essential role for Clp1 in assembly of polyadenylation complex CF IA and Pol II transcription termination

Polyadenylation is a co-transcriptional process that modifies mRNA 3′-ends in eukaryotes. In yeast, CF IA and CPF constitute the core 3′-end maturation complex. CF IA comprises Rna14p, Rna15p, Pcf11p and Clp1p. CF IA interacts with the C-terminal domain of RNA Pol II largest subunit via Pcf11p which links pre-mRNA 3′-end processing to transcription termination. Here, we analysed the role of Clp1p in 3′ processing. Clp1p binds ATP and interacts in CF IA with Pcf11p only. Depletion of Clp1p abolishes transcription termination. Moreover, we found that association of mutations in the ATP-binding domain and in the distant Pcf11p-binding region impair 3′-end processing. Strikingly, these mutations prevent not only Clp1p-Pcf11p interaction but also association of Pcf11p with Rna14p-Rna15p. ChIP experiments showed that Rna15p cross-linking to the 3′-end of a protein-coding gene is perturbed by these mutations whereas Pcf11p is only partially affected. Our study reveals an essential role of Clp1p in CF IA organization. We postulate that Clp1p transmits conformational changes to RNA Pol II through Pcf11p to couple transcription termination and 3′-end processing. These rearrangements likely rely on the correct orientation of ATP within Clp1p

The geological-event reference system, a step towards geological data harmonization

The temporal dimension is an inherent component of geology. In this regard, traditional geological maps can represent a few geological events, yet they hardly account for the entire complex rock history whether sedimentary, crystalline or volcanic. Here, using the RGF research program (French Geological Reference platform) we propose a new methodology based on digital technology and the French historical collection of 1:50 000-scale geological maps. This innovative approach consists of describing, organizing and hierarchizing a series of geological events within a reference framework and linking it to GIS map geometries (polygons, faults, points). In this way, the complete history of geological features can be compiled and stored in digital maps, combining distinct geological events and properties. For a single event, all associated transformations can be represented on maps, facilitating the production of real “palaeo-geological” maps that consider not only traditional sedimentary environments but also possible synchronous weathering, metamorphism, and volcanism. We discuss here an example of French orogenic history. The approach demonstrated here on geological maps can be used with other geological data media (boreholes, seismic reflection profiles, etc.) and thus facilitate a 3D-to-4D scale, with a significant ability to address not only academic community needs, but also themes or issues related to applications required by politics, civil engineering, and society itself, to confront challenges such as natural and anthropic risk reduction and subsurface uses

Etude de la conformation de deux ARN ribosomiques : l'ARN 16S d'Escherichia coli et l'ARN 5S de Xenopus laevis : relations structure-fonction

SIGLECNRS T Bordereau / INIST-CNRS - Institut de l'Information Scientifique et TechniqueFRFranc

Plongée au coeur d\u27une poétique au féminin de la parole martiniquaise

La Poétesse Flaurence BAUDIN retient l\u27attention du public par le biais de belles paroles poétique

Etudes structurales et fonctionnelles de la sous-unité PA du complexe polymérasique du virus influenza

La polymérase du virus de la grippe est un complexe hétérotrimérique composé des sous-unités PB1,PB2 et PA. Le rOle de ce complexe est de répliquer et de transcrire les génomes viraux, au sein du compartiment nucléaire. Malgré les nombreux efforts déployés sur l'étudE de la polymérase, seules des reconstructions par microscopie électronique sont disponibles sur ce complexe. Récemment, notre laboratoire a lancé un projet d'étude structurale et fonctionnelle sur ce complexe, permettant d'obtenir les structures tridimensionnelles dE plusieurs domaines de la sous-unité PB2. Au cours de mes travaux de thése, nous sommes parvenus à résoudre la structure d'un domaine de PA : Le domaine N-terminal contenant 209 résidus. Nous avons montré que ce domaine présente une similarité structurale avec les enzymes de restriction de type Il et qu'il posséde une activité nucléase (DNase et RNase). De ce fait, ce domaine peut être classé dans la grande famille des enzymes à motif PD-D/EXK. Cette étude associée aux données structurales collectées sur la sous unité PB2, permet de mieux comprendre le mécanisme de vol de coiffe, spécifique du virus de la grippe. Enfin, l'obtention de la structure tridimensionnelle de ce domaine présentant une activité enzymatique est prometteur pour la conception rationnelle d'inhibiteur spécifiquE1de l'endonucléase du virus influenza.The influenza virus polymerase is a heterotrimeric complex composed of PA, PB1 and PB2 subunits. This complex is dedicated to replication and transcription of the viral genome, inside the nuclear compartment. Despite much research performed on the polymerase, the only structural data available on this complex are reconstructions made by electronic microscopy. Recently, our laboratory has initiated a structural and functional project on this complex, which led to the structure determination of several PB2 domains. My thesis work led to the structure determination of another domain from the PA subunit: the N-terminal domain encompassing the 209 first residues. We have shown that this domain shares structural similarities with type Il restriction enzymes and that it possesses nuclease activity (DNase and RNase). Indeed, this domain belongs to the PD-D/EXK enzyme family. This work, together with structural data collected on PB2, gives a better view of the cap-snatching mechanism, which is specifie of the influenza virus. Eventually, the availability of the structure of the enzymatically active PA-Nter domain should aid the design of a specifie inhibitor of the influenza virus endonuclease.GRENOBLE1-BU Sciences (384212103) / SudocSudocFranceF

Etudes structurales et fonctionnelles des protéines virales impliquées dans le trafic intracellulaire du génome du virus de la grippe

Le virus de la grippe appartient à la famille des Orthomyxoviridae. Son génome est segmenté en huit ribonucléoprotéines virales (vRNPS) composées chacune d'une molécule d'ARN négatif simple brin recouverte de nucléoprotéines (NP) et associée au complexe de la polymérase. Le virus entre dans la cellule hôte par endocytose, libère dans le cytoplasme ses vRNPs qui gagnent ensuite le noyau cellulaire pour être transcrites et répliquées. Les vRNPs sont importées via l'hétérocomplexe des importines alpha/beta. La NP, composant majoritaire des vRNPs, pourrait être impliquée dans cette étape et serait aussi un candidat idéal dans la mise au point de drogues antivirales. Nous avons voulu déterminer les caractéristiques structurales de NP en complexe avec l'importine alpha 5 humaine. Grâce à la technique de microscopie électronique à transmission, nous avons obtenu un premier modèle à basse résolution du complexe NP/importine alpha. Dans le noyau, les vRNPs sont étroitement liées à la chromatine. En phase tardive du cycle viral, la protéine matricielle M1 se lierait aussi à la chromatine, peut-être pour décrocher les vRNPs. Nous avons pu montrer, par la technique de sédimentation, que les vRNPs se lient aux queues des histones, alors que M1 se fixe sur le domaine globulaire des octamères d'histones. En fin de cycle viral, les vRNPs amplifiées sortent du noyau. Nos tests enzymatiques, d'interaction sur billes et en sédimentation montrent que les protéines virales M1 et NEP servent d'adaptateurs entre les vRNPs et le système d'export nucléaire CRM1/RanGTP. Nous avons aussi obtenu la structure cristallographique du domaine C-terminal de NEP se liant à M1.GRENOBLE1-BU Sciences (384212103) / SudocSudocFranceF