Functional analysis of human T lymphotropic virus type 2 Tax proteins

BACKGROUND: The Tax proteins encoded by human T lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) are transcriptional activators of both the viral long terminal repeat (LTR) and cellular promoters via the CREB and NFkB pathways. In contrast to HTLV-1, HTLV-2 has been classified into four distinct genetic subtypes A, B, C and D defined by phylogenetic analysis of their nucleotide sequences and the size and amino acid sequence of their Tax proteins. In the present study we have analysed and compared the transactivating activities of three Tax 2A and one Tax 2B proteins using LTR and NFkB reporter assays. RESULTS: We found that with the exception of the prototype Tax 2A Mo protein, the other two Tax 2A proteins failed to transactivate either the viral LTR or NFkB promoter in Jurkat and 293T cells. Loss of activity was not associated with either expression levels or an alteration in subcellular distribution as all Tax 2 proteins were predominantly located in the cytoplasm of transfected cells. Analysis of the sequence of the two inactive Tax 2A proteins relative to Mo indicated that one had six amino acid changes and the other had one change in the central region of the protein. Mutations present at the amino and the extreme carboxy termini of Mo resulted in the loss of LTR but not NFkB activation whereas those occurring in the central region of the protein appeared to abolish transactivation of both promoters. Analysis of the transactivation phenotypes of Tax 1, Tax 2A Mo and Tax 2B containing mutations identified in the present study or previously characterised Tax mutations showed that domains required for LTR and NFkB activation are very similar but not identical in all three Tax proteins. CONCLUSION: Our results suggest that loss of activity of two Tax 2A proteins derived from different isolates is associated with multiple amino acid changes relative to Mo in domains required for the activation of the CREB or CREB and NFkB pathways and that these domains are very similar but not identical in Tax 2B and Tax 1. The loss of Tax function in 2A viruses may have implications for their biological and pathogenic properties

Intermolecular masking of the HIV-1 Rev NLS by the cellular protein HIC: Novel insights into the regulation of Rev nuclear import

<p>Abstract</p> <p>Background</p> <p>The HIV-1 regulatory protein Rev, which is essential for viral replication, mediates the nuclear export of unspliced viral transcripts. Rev nuclear function requires active nucleocytoplasmic shuttling, and Rev nuclear import is mediated by the recognition of its Nuclear Localisation Signal (NLS) by multiple import factors, which include transportin and importin β. However, it remains unclear which nuclear import pathway(s) predominate <it>in vivo</it>, and the cellular environment that modulates Rev nucleocytoplasmic shuttling remains to be characterised.</p> <p>Results</p> <p>In our study, we have identified the cellular protein HIC (Human I-mfa domain-Containing protein) as a novel interactor of HIV-1 Rev. We demonstrate that HIC selectively interferes with Rev NLS interaction with importin β and impedes its nuclear import and function, but does not affect Rev nuclear import mediated by transportin. Hence, the molecular determinants mediating Rev-NLS recognition by importin β and transportin appear to be distinct. Furthermore, we have employed HIC and M9 M, a peptide specifically designed to inhibit the transportin-mediated nuclear import pathway, to characterise Rev nuclear import pathways within different cellular environments. Remarkably, we could show that in 293T, HeLa, COS7, Jurkat, U937, THP-1 and CEM cells, Rev nuclear import is cell type specific and alternatively mediated by transportin or importin β, in a mutually exclusive fashion.</p> <p>Conclusions</p> <p>Rev cytoplasmic sequestration by HIC may represent a novel mechanism for the control of Rev function. These studies highlight that the multivalent nature of the Rev NLS for different import receptors enables Rev to adapt its nuclear trafficking strategy.</p

Satisfaction with care among patients with non-metastatic breast cancer: development and first steps of validation of the REPERES-60 questionnaire

<p>Abstract</p> <p>Background</p> <p>The care itinerary for cancer involves difficulties that occur in several different areas, whether in the diagnostic procedures, in surgery, or in adjuvant treatment. The aim of this work was to obtain a valid instrument measuring satisfaction among patients with breast cancer and exploring their care itinerary overall.</p> <p>Methods</p> <p><it>Development phase</it>: Patient focus groups were implemented in two French regions in order to identify areas of satisfaction in relation to the different phases of care provision in breast cancer. On the basis of the literature and the themes and wordings derived from the focus groups, the patients identified several areas of satisfaction, which they found to be partially covered in an American satisfaction measure that has been validated in the French general population (the Consumer Satisfaction Survey in its French version, CSS-VF, 39 items). The patient focus groups suggested adaptation of certain dimensions of this instrument to the potential care providers (37 items) and produced 45 new items in six areas.</p> <p><it>Validation phase</it>: Using a large sample of patients (cohort of 820 women with invasive non-metastatic breast cancer) approached one month after treatment, this phase selected items that were comprehensible (non-response rate < 10%), non-redundant (r < 0.80) and reproducible (test-retest conducted on a sub-sample of 166 patients). The dimensions were identified by factor analysis on the selected items. Divergent and discriminant validity were assessed (relationships with quality of life questionnaire, comparisons between extreme groups).</p> <p>Results</p> <p>Results were in favour of not inserting additional broken-down items into the CSS-VF and retaining 21 new items. The factor analysis found the initial structure of the CSS-VF (39 items in 9 dimensions) and the 21 new items divide up into four dimensions (listening abilities and information provided by doctors, organisation and follow-up of medical care provision, psychological support, material environment). No redundancy was observed between new items and CSS-VF items. Internal consistency was high. Divergent and discriminant validity were satisfactory.</p> <p>Conclusion</p> <p>Adding four new dimensions to the CSS-VF yielded a valid 60-item instrument for assessment of care provided in breast cancer. These promising results now require further investigations of its responsiveness and its robustness in other linguistic, cultural and healthcare settings.</p

Investigating nucleo-cytoplasmic shuttling of the human DEAD-box helicase DDX3

The human DEAD-box helicase DDX3 is a multi-functional protein involved in the regulation of gene expression and additional non-conventional roles as signalling adaptor molecule that are independent of its enzymatic RNA remodeling activity. It is a nucleo-cytoplasmic shuttling protein and it has previously been suggested that dysregulation of its subcellular localization could contribute to tumourigenesis. Indeed, both tumour suppressor and oncogenic functions have been attributed to DDX3. In this study, we investigated the regulation of DDX3's nucleocytoplasmic shuttling. We confirmed that an N-terminal conserved Nuclear Export Signal (NES) is required for export of human DDX3 from the nucleus, and identified three regions within DDX3 that can independently facilitate its nuclear import. We also aimed to identify conditions that alter DDX3's subcellular localisation. Viral infection, cytokine treatment and DNA damage only induced minor changes in DDX3's subcellular distribution as determined by High Content Analysis. However, DDX3's nuclear localization increased in early mitotic cells (during prophase) concomitant with an increase in DDX3 expression levels. Our results are likely to have implications for the proposed use of (nuclear) DDX3 as a prognostic biomarker in cancer

ÉVALUATION DE L'ENSEIGNEMENT AU PREMIER CYCLE DE L'INSA LYON -- Un chantier de taille, qui demeure ouvert

Dans le cadre de la mise en place de l'évaluation des enseignements au premier cycle de l'INSA Lyon, le principal problème a été de permettre un passage à l'échelle de la démarche, tout en respectant le droit des enseignants à la confidentialité des résultats. Il était également nécessaire d'informer les étudiants et les collègues sur l'intérêt de la démarche, tout en sensibilisant dès le départ sur au temps nécessaire. Cependant, des problèmes demeurent et le groupe de travail continue ses réflexions et sa recherche d'outils pour progresser.Dans le cadre de la mise en place de l'évaluation des enseignements au premier cycle de l'INSA Lyon, le principal problème a été de permettre un passage à l'échelle de la démarche, tout en respectant le droit des enseignants à la confidentialité des résultats. Il était également nécessaire d'informer les étudiants et les collègues sur l'intérêt de la démarche, tout en sensibilisant dès le départ sur au temps nécessaire. Cependant, des problèmes demeurent et le groupe de travail continue ses réflexions et sa recherche d'outils pour progresser

Identifying roles of the scion and the rootstock in regulating plant development and functioning under different phosphorus supplies in grapevine

COntinental To coastal Ecosystems: evolution, adaptability and governanc

Reversible transitions between noradrenergic and mesenchymal tumor identities define cell plasticity in neuroblastoma

Noradrenergic and mesenchymal identities have been characterized in neuroblastoma cell lines according to their epigenetic landscapes and core regulatory circuitries. However, their relationship and relative contribution in patient tumors remain poorly defined. We now document spontaneous and reversible plasticity between the two identities, associated with epigenetic reprogramming, in several neuroblastoma models. Interestingly, xenografts with cells from each identity eventually harbor a noradrenergic phenotype suggesting that the microenvironment provides a powerful pressure towards this phenotype. Accordingly, such a noradrenergic cell identity is systematically observed in single-cell RNA-seq of 18 tumor biopsies and 15 PDX models. Yet, a subpopulation of these noradrenergic tumor cells presents with mesenchymal features that are shared with plasticity models, indicating that the plasticity described in these models has relevance in neuroblastoma patients. This work therefore emphasizes that intrinsic plasticity properties of neuroblastoma cells are dependent upon external cues of the environment to drive cell identity

L’épave de Port Berteau II (Charente-Maritime)

Découverte en 1973 dans la Charente, en aval de Saintes et par 7 m de fond, l’épave de Port Berteau II a été fouillée de 1992 à 1997 avec une double problématique reposant sur la mise en relation des vestiges de la coque avec son environnement.Conduite selon les techniques classiques de l’archéologie subaquatique, l’étude de l’épave, de l’étrave à l’étambot, a permis de dresser le -portrait architectural » d’un caboteur du début du Vile s. Long de 1430 m et bâti à franc-bord embrure première », ce bateau a été construit pour la navigation fluvio-maritime et pouvait transporter une cargaison d’une dizaine de tonnes. L’analyse du milieu fluvial, relevant de ce que l’on pourrait qualifier d’archéologie nautique, a abouti quant à elle à une restitution du paysage naturel contemporain de l’épave. Le caboteur de Port Berteau II est désormais une référence pour l’histoire de la construction à carvel dans le Ponant uniquement attestée jusqu’alors par des textes du XVe s.Discovered in 1973, the Port Berteau wreck lies at a depth of 7 metres in the Charente river below Samtes. Excavations (1992-1997) were carried out with the dual objective of studying both the hull’s remains and its natural environment.Using the traditional techniques of ship archaeology to examine the wreck from the stem to the stern-post has enabled archaeologists to draw an ‘architectural portrait" of an early 7th century coaster. The 14.30 metre carvel built "frame-first" boat was intended for river navigation and built to carry a cargo of up to ten tonnes. Using what could be called a naval archaeology approach, researchers have analysed the nver environment to reconstruct the natural landscape of the period. The Port Berteau II coaster thus provides a reference for carvel construction history in the Ponant, reviously attested only in 15th century texts

In vitro nuclear interactome of the HIV-1 Tat protein

<p>Abstract</p> <p>Background</p> <p>One facet of the complexity underlying the biology of HIV-1 resides not only in its limited number of viral proteins, but in the extensive repertoire of cellular proteins they interact with and their higher-order assembly. HIV-1 encodes the regulatory protein Tat (86–101aa), which is essential for HIV-1 replication and primarily orchestrates HIV-1 provirus transcriptional regulation. Previous studies have demonstrated that Tat function is highly dependent on specific interactions with a range of cellular proteins. However they can only partially account for the intricate molecular mechanisms underlying the dynamics of proviral gene expression. To obtain a comprehensive nuclear interaction map of Tat in T-cells, we have designed a proteomic strategy based on affinity chromatography coupled with mass spectrometry.</p> <p>Results</p> <p>Our approach resulted in the identification of a total of 183 candidates as Tat nuclear partners, 90% of which have not been previously characterised. Subsequently we applied <it>in silico </it>analysis, to validate and characterise our dataset which revealed that the Tat nuclear interactome exhibits unique signature(s). First, motif composition analysis highlighted that our dataset is enriched for domains mediating protein, RNA and DNA interactions, and helicase and ATPase activities. Secondly, functional classification and network reconstruction clearly depicted Tat as a polyvalent protein adaptor and positioned Tat at the nexus of a densely interconnected interaction network involved in a range of biological processes which included gene expression regulation, RNA biogenesis, chromatin structure, chromosome organisation, DNA replication and nuclear architecture.</p> <p>Conclusion</p> <p>We have completed the <it>in vitro </it>Tat nuclear interactome and have highlighted its modular network properties and particularly those involved in the coordination of gene expression by Tat. Ultimately, the highly specialised set of molecular interactions identified will provide a framework to further advance our understanding of the mechanisms of HIV-1 proviral gene silencing and activation.</p