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I2BC Publications

I2BC is ranked in the Nature Index 2017-2018


  • F. Acosta, J. Agapito, A. M. Cabibbe, T. Cáceres, C. Sola, L. Pérez-Lago, E. Abascal, M. Herranz, E. Meza, B. Klotoe, P. Muñoz, G. M. Rossolini, A. Bartoloni, E. Tortoli, D. M. Cirillo, E. Gotuzzo, et D. García de Viedma, « Exportation of MDR TB to Europe from Setting with Actively Transmitted Persistent Strains in Peru », Emerging Infectious Diseases, vol. 25, nᵒ 3, p. 596-598, mars 2019.
    Résumé : We performed a cross-border molecular epidemiology analysis of multidrug-resistant tuberculosis in Peru, Spain, and Italy. This analysis revealed frequent transmission in Peru and exportation of a strain that recreated similar levels of transmission in Europe during 2007-2017. Transnational efforts are needed to control transmission of multidrug-resistant tuberculosis globally.
    Mots-clés : antimicrobial resistance, bacteria, Europe, exportation, Florence, IGEPE, Italy, Lima, Madrid, MDR, MICROBIO, migration, MIRU-VNTR, molecular epidemiology, mycobacterium, Mycobacterium tuberculosis, Peru, single-nucleotide polymorphism, Spain, spoligotype, transmission, tuberculosis, tuberculosis and other mycobacteria.

  • G. Agbota, N. Fievet, B. Heude, M. Accrombessi, U. Ahouayito, E. Yovo, D. Dossa, L. Dramane, A. Gartner, S. Ezinmègnon, J. Yugueros Marcos, L. Vachot, P. Tissières, A. Massougbodji, Y. Martin-Prével, M. Cot, et V. Briand, « Poor maternal anthropometric status before conception is associated with a deleterious infant growth during the first year of life: a longitudinal preconceptional cohort », Pediatric Obesity, août 2019.
    Résumé : BACKGROUND: According to the Developmental Origins of Health and Diseases concept, exposures in the preconception period may be critical. For the first time, we evaluated the effect of preconception poor anthropometric status on infant's growth in sub-Saharan Africa. METHODS: A mother-child cohort was followed prospectively from preconception to 1 year old in Benin. Maternal anthropometric status was assessed by prepregnancy body mass index (BMI), approximated by BMI at the first antenatal visit before 7 weeks' gestation, and gestational weight gain (GWG). BMI was categorized as underweight, normal, overweight, and obesity according to World Health Organization standards. GWG was categorized as low (<7 kg), mild (7-12 kg), and high (>12 kg). In infant, stunting and wasting were defined as length-for-age and weight-for-length z scores less than -2 SD, respectively. We evaluated the association between BMI/GWG and infant's weight and length at birth and during the first year of life, as well as with stunting and wasting at 12 months using mixed linear and logistic regression models. RESULTS: In multivariate, preconceptional underweight was associated with a lower infant's weight at birth and during the first year (-164 g; 95% CI, -307 to -22; and -342 g; 95% CI, -624 to -61, respectively) and with a higher risk of stunting at 12 months (adjusted odds ratio [aOR] = 3.98; 95% CI, 1.01-15.85). Furthermore, preconceptional obesity and a high GWG were associated with a higher weight and length at birth and during the first year. CONCLUSION: Underweight and obesity before conception as well as GWG were associated with infant's growth. These results argue for preventive interventions starting as early as the preconception period to support child long-term health.
    Mots-clés : ESHR, infant growth, maternal anthropometric status, MICROBIO, Preconceptional cohort study, pregnancy.

  • G. Agbota, K. Polman, F. T. Wieringa, M. Campos-Ponce, M. Accrombessi, E. Yovo, C. Roucher, S. Ezinmègnon, J. Y. Marcos, L. Vachot, P. Tissières, A. Massougbodji, N. Fievet, M. Cot, et V. Briand, « Maternal malaria but not schistosomiasis is associated with a higher risk of febrile infection in infant during the first 3 months of life: A mother-child cohort in Benin », PloS One, vol. 14, nᵒ 9, p. e0222864, 2019.
    Résumé : BACKGROUND: Malaria and schistosomiasis represent two of the most prevalent and disabling parasitic infections in developing countries. Few studies have evaluated the effect of maternal schistosomiasis and malaria in the peri-conceptional period on infant's risk of infection. METHODS: In Benin, women were followed from the preconception period until delivery. Subsequently, their children were followed from birth to 3 months of age. Pre-pregnancy malaria, malaria in pregnancy (MiP)-determined monthly using a thick blood smear-and urinary schistosomiasis-determined once before pregnancy and once at delivery using urine filtration-were the main maternal exposures. Infant's febrile infection (fever with respiratory, gastrointestinal and/or cutaneous clinical signs anytime during follow-up) was the main outcome. In a secondary analysis, we checked the relation of malaria and schistosomiasis with infant's hemoglobin (Hb) concentration. Both effects were separately assessed using logistic/mixed linear regression models. RESULTS: The prevalence of MiP was 35.7% with 10.8% occurring during the 1st trimester, and the prevalence of schistosomiasis was 21.8%. From birth to 3 months, 25.3% of infants had at least one episode of febrile infection. In multivariate analysis, MiP, particularly malaria in the 1st trimester, was significantly associated with a higher risk of infant's febrile infection (aOR = 4.99 [1.1; 22.6], p = 0.03). In secondary results, pre-pregnancy malaria and schistosomiasis were significantly associated with a lower infant's Hb concentration during the first 3 months. CONCLUSION: We evidenced the deleterious effect of maternal parasitic infections on infant's health. Our results argue in favor of the implementation of preventive strategies as early as in the peri-conception.
    Mots-clés : ESHR, MICROBIO.

  • A. Alard, C. Marboeuf, B. Fabre, C. Jean, Y. Martineau, F. Lopez, P. Vende, D. Poncet, R. J. Schneider, C. Bousquet, et S. Pyronnet, « Differential Regulation of the Three Eukaryotic mRNA Translation Initiation Factor (eIF) 4Gs by the Proteasome », Frontiers in Genetics, vol. 10, p. 254, mars 2019.
    Résumé : The 4G family of eukaryotic mRNA translation initiation factors is composed of three members (eIF4GI, eIF4GII, and DAP5). Their specific roles in translation initiation are under intense investigations, but how their respective intracellular amounts are controlled remains poorly understood. Here we show that eIF4GI and eIF4GII exhibit much shorter half-lives than that of DAP5. Both eIF4GI and eIF4GII proteins, but not DAP5, contain computer-predicted PEST motifs in their N-termini conserved across the animal kingdom. They are both sensitive to degradation by the proteasome. Under normal conditions, eIF4GI and eIF4GII are protected from proteasomal destruction through binding to the detoxifying enzyme NQO1 [NAD(P)H:quinone oxidoreductase]. However, when cells are exposed to oxidative stress both eIF4GI and eIF4GII, but not DAP5, are degraded by the proteasome in an N-terminal-dependent manner, and cell viability is more compromised upon silencing of DAP5. These findings indicate that the three eIF4G proteins are differentially regulated by the proteasome and that persistent DAP5 plays a role in cell survival upon oxidative stress.
    Mots-clés : 20s proteasome, DAP5, NQO1, Nrf2, Oxidative Stress, Pest, Proteasome, ROTA, VIRO.
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  • W. R. Algar, N. Hildebrandt, S. S. Vogel, et I. L. Medintz, « FRET as a biomolecular research tool - understanding its potential while avoiding pitfalls », Nature Methods, vol. 16, nᵒ 9, p. 815-829, sept. 2019.
    Résumé : The applications of Förster resonance energy transfer (FRET) grow with each year. However, different FRET techniques are not applied consistently, nor are results uniformly presented, which makes implementing and reproducing FRET experiments challenging. We discuss important considerations for designing and evaluating ensemble FRET experiments. Alongside a primer on FRET basics, we provide guidelines for making experimental design choices such as the donor-acceptor pair, instrumentation and labeling chemistries; selecting control experiments to unambiguously demonstrate FRET and validate that the experiments provide meaningful data about the biomolecular process in question; analyzing raw data and assessing the results; and reporting data and experimental details in a manner that easily allows for reproducibility. Some considerations are also given for FRET assays and FRET imaging, especially with fluorescent proteins. Our goal is to motivate and empower all biologists to consider FRET for the powerful research tool it can be.
    Mots-clés : B3S, NANO.

  • A. Aliouat, I. Hatin, P. Bertin, P. François, V. Stierlé, O. Namy, S. Salhi, et O. Jean-Jean, « Divergent effects of translation termination factor eRF3A and nonsense-mediated mRNA decay factor UPF1 on the expression of uORF carrying mRNAs and ribosome protein genes », RNA biology, p. 1-13, oct. 2019.
    Résumé : In addition to its role in translation termination, eRF3A has been implicated in the nonsense-mediated mRNA decay (NMD) pathway through its interaction with UPF1. NMD is a RNA quality control mechanism, which detects and degrades aberrant mRNAs as well as some normal transcripts including those that harbour upstream open reading frames in their 5' leader sequence. In this study, we used RNA-sequencing and ribosome profiling to perform a genome wide analysis of the effect of either eRF3A or UPF1 depletion in human cells. Our bioinformatics analyses allow to delineate the features of the transcripts controlled by eRF3A and UPF1 and to compare the effect of each of these factors on gene expression. We find that eRF3A and UPF1 have very different impacts on the human transcriptome, less than 250 transcripts being targeted by both factors. We show that eRF3A depletion globally derepresses the expression of mRNAs containing translated uORFs while UPF1 knockdown derepresses only the mRNAs harbouring uORFs with an AUG codon in an optimal context for translation initiation. Finally, we also find that eRF3A and UPF1 have opposite effects on ribosome protein gene expression. Together, our results provide important elements for understanding the impact of translation termination and NMD on the human transcriptome and reveal novel determinants of ribosome biogenesis regulation.
    Mots-clés : DBG, eRF3, GSPT1, GST, nonsense-mediated mRNA decay, ribosome protein genes, translation termination, uORF, UPF1.

  • I. Altinoglu, C. J. Merrifield, et Y. Yamaichi, « Single molecule super-resolution imaging of bacterial cell pole proteins with high-throughput quantitative analysis pipeline », Scientific Reports, vol. 9, nᵒ 1, p. 6680, avr. 2019.
    Résumé : Bacteria show sophisticated control of their cellular organization, and many bacteria deploy different polar landmark proteins to organize the cell pole. Super-resolution microscopy, such as Photo-Activated Localization Microscopy (PALM), provides the nanoscale localization of molecules and is crucial for better understanding of organization and dynamics in single-molecule. However, analytical tools are not fully available yet, in particular for bacterial cell biology. For example, quantitative and statistical analyses of subcellular localization with multiple cells from multiple fields of view are lacking. Furthermore, brightfield images are not sufficient to get accurate contours of small and low contrast bacterial cells, compared to subpixel presentation of target molecules. Here we describe a novel analytic tool for PALM which integrates precisely drawn cell outlines, of either inner membrane or periplasm, labelled by PALM-compatible fluorescent protein fusions, with molecule data for >10,000 molecules from >100 cells by fitting each cell into an oval arc. In the vibrioid bacterium Vibrio cholerae, the polar anchor HubP constitutes a big polar complex which includes multiple proteins involved in chemotaxis and the flagellum. With this pipeline, HubP is shown to be slightly skewed towards the inner curvature side of the cell, while its interaction partners showed rather loose polar localization.
    Mots-clés : BIOCELL, DBG, EQYY.

  • A. Arfaoui, C. Rioualen, V. Azzoni, G. Pinna, P. Finetti, J. Wicinski, E. Josselin, M. Macario, R. Castellano, C. Léonard-Stumpf, A. Bal, A. Gros, S. Lossy, M. Kharrat, Y. Collette, F. Bertucci, D. Birnbaum, H. Douik, G. Bidaut, E. Charafe-Jauffret, et C. Ginestier, « A genome-wide RNAi screen reveals essential therapeutic targets of breast cancer stem cells », EMBO molecular medicine, p. e9930, sept. 2019.
    Résumé : Therapeutic resistance is a major clinical challenge in oncology. Evidence identifies cancer stem cells (CSCs) as a driver of tumor evolution. Accordingly, the key stemness property unique to CSCs may represent a reservoir of therapeutic target to improve cancer treatment. Here, we carried out a genome-wide RNA interference screen to identify genes that regulate breast CSCs-fate (bCSC). Using an interactome/regulome analysis, we integrated screen results in a functional mapping of the CSC-related processes. This network analysis uncovered potential therapeutic targets controlling bCSC-fate. We tested a panel of 15 compounds targeting these regulators. We showed that mifepristone, salinomycin, and JQ1 represent the best anti-bCSC activity. A combination assay revealed a synergistic interaction of salinomycin/JQ1 association to deplete the bCSC population. Treatment of primary breast cancer xenografts with this combination reduced the tumor-initiating cell population and limited metastatic development. The clinical relevance of our findings was reinforced by an association between the expression of the bCSC-related networks and patient prognosis. Targeting bCSCs with salinomycin/JQ1 combination provides the basis for a new therapeutic approach in the treatment of breast cancer.
    Mots-clés : breast cancer, cancer stem cells, JQ1, PARI, PF, RNAi screen, salinomycin.

  • C. Arrondel, S. Missoury, R. Snoek, J. Patat, G. Menara, B. Collinet, D. Liger, D. Durand, O. Gribouval, O. Boyer, L. Buscara, G. Martin, E. Machuca, F. Nevo, E. Lescop, D. A. Braun, A. - C. Boschat, S. Sanquer, I. C. Guerrera, P. Revy, M. Parisot, C. Masson, N. Boddaert, M. Charbit, S. Decramer, R. Novo, M. - A. Macher, B. Ranchin, J. Bacchetta, A. Laurent, S. Collardeau-Frachon, A. M. van Eerde, F. Hildebrandt, D. Magen, C. Antignac, H. van Tilbeurgh, et G. Mollet, « Defects in t6A tRNA modification due to GON7 and YRDC mutations lead to Galloway-Mowat syndrome », Nature Communications, vol. 10, nᵒ 1, p. 3967, sept. 2019.
    Résumé : N6-threonyl-carbamoylation of adenosine 37 of ANN-type tRNAs (t6A) is a universal modification essential for translational accuracy and efficiency. The t6A pathway uses two sequentially acting enzymes, YRDC and OSGEP, the latter being a subunit of the multiprotein KEOPS complex. We recently identified mutations in genes encoding four out of the five KEOPS subunits in children with Galloway-Mowat syndrome (GAMOS), a clinically heterogeneous autosomal recessive disease characterized by early-onset steroid-resistant nephrotic syndrome and microcephaly. Here we show that mutations in YRDC cause an extremely severe form of GAMOS whereas mutations in GON7, encoding the fifth KEOPS subunit, lead to a milder form of the disease. The crystal structure of the GON7/LAGE3/OSGEP subcomplex shows that the intrinsically disordered GON7 protein becomes partially structured upon binding to LAGE3. The structure and cellular characterization of GON7 suggest its involvement in the cellular stability and quaternary arrangement of the KEOPS complex.
    Mots-clés : B3S, FAAM.
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  • A. - A. Arteni, A. M. LaFountain, M. T. A. Alexandre, M. Fradot, M. M. Mendes-Pinto, J. - A. Sahel, S. Picaud, H. A. Frank, B. Robert, et A. A. Pascal, « Carotenoid composition and conformation in retinal oil droplets of the domestic chicken », PloS One, vol. 14, nᵒ 5, p. e0217418, 2019.
    Résumé : Carotenoid-containing oil droplets in the avian retina act as cut-off filters to enhance colour discrimination. We report a confocal resonance Raman investigation of the oil droplets of the domestic chicken, Gallus gallus domesticus. We show that all carotenoids present are in a constrained conformation, implying a locus in specific lipid binding sites. In addition, we provide proof of a recent conclusion that all carotenoid-containing droplets contain a mixture of all carotenoids present, rather than only a subset of them-a conclusion that diverges from the previously-held view. Our results have implications for the mechanism(s) giving rise to these carotenoid mixtures in the differently-coloured droplets.
    Mots-clés : B3S, LBMS.

  • J. Astier, A. Mounier, J. Santolini, S. Jeandroz, et D. Wendehenne, « The evolution of nitric oxide signalling diverges between the animal and the green lineages », Journal of Experimental Botany, mars 2019.
    Résumé : Nitric oxide (NO) is a ubiquitous signalling molecule with widespread distribution in prokaryotes and eukaryotes where it is involved in countless physiological processes. While the mechanisms governing NO synthesis and signalling are well established in animals, the situation is less clear in the green lineage. Recent investigations have shown that NO synthase (NOS), the major enzymatic source for NO in animals, is absent in land plants but present in a limited number of algae. First detailed analysis highlighted that these new NOSs are functional but display specific structural features and probably original catalytic activities. Completing this picture, analyses were undertaken in order to investigate whether major components of the prototypic NO/cyclic GMP signalling cascades mediating many physiological effects of NO in animals were also present in plants. Only few homologues of soluble guanylate cyclases, cGMP-dependent protein kinases, cyclic nucleotide-gated channels and cGMP-regulated phosphodiesterases, were identified in some algal species and their presence did not correlate with that of NOSs. In contrast, GSNO reductase, a critical regulator of S-nitrosothiols, was recurrently found. Overall, these findings highlight that plants do not mediate NO signalling through the classical NO/cGMP-signalling module and support the concept that S-nitrosation is a ubiquitous NO-dependent signalling mechanism.
    Mots-clés : Algae, B3S, cGMP, cGMP-dependent protein kinase, cyclic nucleotide-gated channel, Guanylate cyclase, LSOD, Nitric oxide, Nitric oxide synthase, phosphodiesterase, Plant, Signalling.

  • C. Aubry, J. - L. Pernodet, et S. Lautru, « A set of modular and integrative vectors for synthetic biology in Streptomyces », Applied and Environmental Microbiology, vol. 85, nᵒ 16, juin 2019.
    Résumé : With the development of synthetic biology in the field of (actinobacteria) specialized metabolism, new tools are needed for the design or refactoring of biosynthetic gene clusters. If libraries of synthetic parts (such as promoters or ribosome binding sites) and DNA cloning methods have been developed, to our knowledge, not many vectors designed for the flexible cloning of biosynthetic gene clusters have been constructed.We report here the construction of a set of 12 standardized and modular vectors designed to afford the construction or the refactoring of biosynthetic gene clusters in Streptomyces species, using a large panel of cloning methods. Three different resistance cassettes and four orthogonal integration systems are proposed. In addition, FRT sites were incorporated to allow the recycling of antibiotic markers and to limit the risks of unwanted homologous recombination in Streptomyces strains, when several vectors are used. The functionality and proper integration of the vectors in three commonly used Streptomyces strains, as well as the functionality of the Flp-catalysed excision were all confirmed.To illustrate some possible uses of our vectors, we refactored the albonoursin gene cluster from Streptomyces noursei using the Biocrick assembly method. We also used the seamless Ligase Chain Reaction cloning method to assemble a transcription unit in one of the vectors and genetically complement a mutant strain.IMPORTANCE One of the strategies employed today to obtain new bioactive molecules with potential applications for human health (for example antimicrobial or anticancer agents) is synthetic biology. Synthetic biology is used to biosynthesize new unnatural specialized metabolites, or to force the expression of otherwise silent natural biosynthetic gene clusters. To assist the development of synthetic biology in the field of specialized metabolism, we constructed and are offering to the community a set of vectors that were intended to facilitate DNA assembly and integration in actinobacteria chromosome. These vectors are compatible with various DNA cloning and assembling methods. They are standardized and modular, allowing the easy exchange of a module by another one of the same nature. Although designed for the assembly or the refactoring of specialized metabolite gene clusters, they have a broader potential utility, for protein production or genetic complementation, for example.
    Mots-clés : ACTINO, MICROBIO.

  • C. Badel, G. Erauso, A. Gomez, R. Catchpole, M. Gonnet, J. Oberto, P. Forterre, et V. Da Cunha, « The global distribution and evolutionary history of the pT26-2 archaeal plasmid family », Environmental Microbiology, sept. 2019.
    Résumé : Although plasmids play an important role in biological evolution, the number of plasmid families well characterized in terms of geographical distribution and evolution remains limited, especially in Archaea. Here, we describe the first systematic study of an archaeal plasmid family, the pT26-2 plasmid family. The in-depth analysis of the distribution, biogeography and host-plasmid co-evolution patterns of 26 integrated and 3 extrachromosomal plasmids of this plasmid family shows that they are widespread in Thermococcales and Methanococcales isolated from around the globe but are restricted to these two orders. All members of the family share 7 core genes but employ different integration and replication strategies. Phylogenetic analysis of the core genes and CRISPR spacer distribution suggest that plasmids of the pT26-2 family evolved with their hosts independently in Thermococcales and Methanococcales, despite these hosts exhibiting similar geographic distribution. Remarkably, core genes are conserved even in integrated plasmids that have lost replication genes and/or replication origins suggesting that they may be beneficial for their hosts. We hypothesise that the core proteins encode for a novel type of DNA/protein transfer mechanism, explaining the widespread oceanic distribution of the pT26-2 plasmid family. This article is protected by copyright. All rights reserved.
    Mots-clés : ARCHEE, MICROBIO.

  • C. Badel, V. Da Cunha, R. Catchpole, P. Forterre, et J. Oberto, « WASPS: Web-Assisted Symbolic Plasmid Synteny Server », Bioinformatics (Oxford, England), oct. 2019.
    Résumé : MOTIVATION: Comparative plasmid genome analyses require complex tools, the manipulation of large numbers of sequences and constitute a daunting task for the wet bench experimentalist. Dedicated plasmid databases are sparse, only comprise bacterial plasmids and provide exclusively access to sequence similarity searches. RESULTS: We have developed WASPS (Web-Assisted Symbolic Plasmid Synteny), a web service granting protein and DNA sequence similarity searches against a database comprising all completely sequenced natural plasmids from bacterial, archaeal and eukaryal origin. This database pre-calculates orthologous protein clustering and enables WASPS to generate fully resolved plasmid synteny maps in real time using internal and user-provided DNA sequences. AVAILABILITY AND IMPLEMENTATION: WASPS queries befit all current browsers such as Firefox, Edge or Safari while the best functionality is achieved with Chrome. Internet Explorer is not supported. WASPS is freely accessible at SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

  • M. Bakail, A. Gaubert, J. Andreani, G. Moal, G. Pinna, E. Boyarchuk, M. - C. Gaillard, R. Courbeyrette, C. Mann, J. - Y. Thuret, B. Guichard, B. Murciano, N. Richet, A. Poitou, C. Frederic, M. - H. Le Du, M. Agez, C. Roelants, Z. A. Gurard-Levin, G. Almouzni, N. Cherradi, R. Guerois, et F. Ochsenbein, « Design on a Rational Basis of High-Affinity Peptides Inhibiting the Histone Chaperone ASF1 », Cell Chemical Biology, sept. 2019.
    Résumé : Anti-silencing function 1 (ASF1) is a conserved H3-H4 histone chaperone involved in histone dynamics during replication, transcription, and DNA repair. Overexpressed in proliferating tissues including many tumors, ASF1 has emerged as a promising therapeutic target. Here, we combine structural, computational, and biochemical approaches to design peptides that inhibit the ASF1-histone interaction. Starting from the structure of the human ASF1-histone complex, we developed a rational design strategy combining epitope tethering and optimization of interface contacts to identify a potent peptide inhibitor with a dissociation constant of 3 nM. When introduced into cultured cells, the inhibitors impair cell proliferation, perturb cell-cycle progression, and reduce cell migration and invasion in a manner commensurate with their affinity for ASF1. Finally, we find that direct injection of the most potent ASF1 peptide inhibitor in mouse allografts reduces tumor growth. Our results open new avenues to use ASF1 inhibitors as promising leads for cancer therapy.
    Mots-clés : AMIG, B3S, Cancer, Cell Penetrating Peptide, Chromatin, DBG, Drug Design, Epigenetics, INTGEN, PARI, Peptide Inhibitor, PF, Protein Binding, Protein-Protein Interaction, Rosetta Design, SEN, X-Ray Crystallography.

  • M. Bakail, S. Rodriguez-Marin, Z. Hegedues, M. E. Perrin, F. Ochsenbein, et A. J. Wilson, « Recognition of ASF1 by Using Hydrocarbon-Constrained Peptides », Chembiochem, vol. 20, nᵒ 7, p. 891-895, avr. 2019.
    Résumé : Inhibiting the histone H3-ASF1 (anti-silencing function 1) protein-protein interaction (PPI) represents a potential approach for treating numerous cancers. As an alpha-helix-mediated PPI, constraining the key histone H3 helix (residues 118-135) is a strategy through which chemical probes might be elaborated to test this hypothesis. In this work, variant H3(118-135) peptides bearing pentenylglycine residues at the i and i+4 positions were constrained by olefin metathesis. Biophysical analyses revealed that promotion of a bioactive helical conformation depends on the position at which the constraint is introduced, but that the potency of binding towards ASF1 is unaffected by the constraint and instead that enthalpy-entropy compensation occurs.
    Mots-clés : AMIG, B3S, chemical biology, complex, constrained peptides, helix, histone chaperones, modulators, protein surface recognition, protein-protein interactions histone chaperonnes constrained peptides protein surface recognition chemical biology, protein-protein interactions, replication, stapled peptides, structural basis.

  • A. Baroin-Tourancheau, Y. Jaszczyszyn, X. Benigni, et L. Amar, « Evaluating and Correcting Inherent Bias of microRNA Expression in Illumina Sequencing Analysis », Frontiers in Molecular Biosciences, vol. 6, p. 17, 2019.
    Résumé : microRNA (miRNA) expression profiles based on the highly powerful Illumina sequencing technology rely on the construction of cDNA libraries in which adaptor ligation is known to deeply favor some miRNAs over others. This introduces erroneous measurements of the miRNA abundances and relative miRNA quantities in biological samples. Here, by using the commercial miRXplore Universal Reference that contains an equimolar mixture of 963 animal miRNAs and TruSeq or bulged adaptors, we describe a method for correcting ligation biases in expression profiles obtained with standard protocols of cDNA library construction and provide data for quantifying the true miRNA abundances in biological samples. Ligation biases were evaluated at three ratios of miRNA to 3'-adaptor and four numbers of polymerase chain reaction amplification cycles by calculating efficiency captures/correcting factors for each miRNA. We show that ligation biases lead to over- or under-expression covering a 105 amplitude range. We also show that, at each miRNA:3'-adaptor ratio, coefficients of variation (CVs) of efficiency captures calculated over the four number of amplification cycles using sliding windows of 10 values ranged from 0.1 for the miRNAs of high expression to 0.6 for the miRNAs of low expression. Efficiency captures of miRNAs of high and low expression in profiles are therefore differently impacted by the number of amplification cycles. Importantly, we observed that at a given number of amplification cycles, CVs of efficiency captures calculated over the three miRNA:3'-adaptor ratios displayed a steady value of 0.3 +/- 0.05 STD for miRNAs of high and low expression. This allows, at a given number of amplification cycles, accurate comparison of miRNA expression between biological samples over a substantial expression range. Finally we provide tables of correcting factors that allow to measure the abundances of 963 miRNAs in biological samples from TruSeq-based expression profiles and, an example of their use by characterizing miRNAs of the let-7, miR-26, miR-29, and miR-30 families as the more abundant miRNAs of the rat adult cerebellum.
    Mots-clés : cerebellum, high-throughput sequencing, Illumina technology, ligation bias, miRNA abundance, miRNA expression profile, NGS, PF.

  • H. Barreteau, M. Vandervennet, L. Guedon, V. Point, S. Canaan, S. Rebuffat, J. Peduzzi, et A. Carre-Mlouka, « Haloarcula sebkhae sp. nov., an extremely halophilic archaeon from Algerian hypersaline environment », International Journal of Systematic and Evolutionary Microbiology, vol. 69, nᵒ 3, p. 732-738, mars 2019.
    Résumé : A halophilic organism, SWO25(T) , was isolated from water sampled in Algeria at the salt lake (sebkha) of Ouargla. The novel strain stained Gram-negative, and cells were pleomorphic with a red pigmentation. Strain SWO25(T) - grew optimally at 35-45 degrees C, at pH 6.0-8.0 and 0.05-0.25 M MgCl2 concentrations. Cells were extremely halophilic, with optimal growth at 4.3-5.1 M NaCl. The predominant membrane polar lipids were C20C20 glycerol diether derivatives of phosphatidylglycerol, phosphatidylglycerol phosphate, phosphatidylglycerol sulfate, triglycosyl diether and diglycosyl diether. The major respiratory menaquinone component was MK-8. Cells were highly tolerant to the presence of decane and isooctane in the growth medium. Chemotaxonomic properties supported the assignment of strain SWO25(T) to the genus Haloarcula. The DNA G+C content was 61.1 mol%. DNA-DNA hybridization and phylogenetic analyses of the 16S rRNA and rpoB' genes showed that strain SWO25(T) is distinct from known Haloarcula species. Based on phenotypic, chemotaxonomic, genotypic and phylogenetic data, we describe a novel species of the genus Haloarcula, for which the name Haloarcula sebkhae sp. nov. is proposed. The type strain is SWO25(T)(=CIP 110583(T) =JCM 19018(T)).
    Mots-clés : 16s ribosomal-rna, dna hybridization, ENVBAC, gen. nov., genera, haloarchaea, Haloarcula, halophilic archaeon, heterogeneity, hypersaline environments, MICROBIO, mukohataei, organic-solvent tolerance, rapid method, salt lake, sebkha, sequence.

  • E. M. Bayer, T. Calì, F. Giordano, A. Hamacher-Brady, et L. Pellegrini, « EMBO Workshop: Membrane Contact Sites in Health and Disease », Contact, vol. 2, p. 2515256419825931, janv. 2019.

  • P. Béguin, Y. Chekli, G. Sezonov, P. Forterre, et M. Krupovic, « Sequence motifs recognized by the casposon integrase of Aciduliprofundum boonei », Nucleic Acids Research, mai 2019.
    Résumé : Casposons are a group of bacterial and archaeal DNA transposons encoding a specific integrase, termed casposase, which is homologous to the Cas1 enzyme responsible for the integration of new spacers into CRISPR loci. Here, we characterized the sequence motifs recognized by the casposase from a thermophilic archaeon Aciduliprofundum boonei. We identified a stretch of residues, located in the leader region upstream of the actual integration site, whose deletion or mutagenesis impaired the concerted integration reaction. However, deletions of two-thirds of the target site were fully functional. Various single-stranded 6-FAM-labelled oligonucleotides derived from casposon terminal inverted repeats were as efficiently incorporated as duplexes into the target site. This result suggests that, as in the case of spacer insertion by the CRISPR Cas1-Cas2 integrase, casposon integration involves splaying of the casposon termini, with single-stranded ends being the actual substrates. The sequence critical for incorporation was limited to the five terminal residues derived from the 3' end of the casposon. Furthermore, we characterize the casposase from Nitrosopumilus koreensis, a marine member of the phylum Thaumarchaeota, and show that it shares similar properties with the A. boonei enzyme, despite belonging to a different family. These findings further reinforce the mechanistic similarities and evolutionary connection between the casposons and the adaptation module of the CRISPR-Cas systems.
    Mots-clés : ARCHEE, MICROBIO.

  • L. Belot, A. Albertini, et Y. Gaudin, « Structural and cellular biology of rhabdovirus entry », Advances in Virus Research, vol. 104, p. 147-183, 2019.
    Résumé : Rhabdoviruses are enveloped viruses with a negative-sense single strand RNA genome and are widespread among a great variety of organisms. In their membrane, they have a single glycoprotein (G) that mediates both virus attachment to cellular receptors and fusion between viral and endosomal membranes allowing viral genome release in the cytoplasm. We present structural and cellular aspects of Rhabdovirus entry into their host cell with a focus on vesicular stomatitis virus (VSV) and rabies virus (RABV) for which the early events of the viral cycle have been extensively studied. Recent data have shown that the only VSV receptors are the members of the LDL-R family. This is in contrast with RABV for which multiple receptors belonging to unrelated families have been identified. Despite having different receptors, after attachment, rhabdovirus internalization occurs through clathrin-mediated endocytosis (CME) in an actin-dependent manner. There are still debates about the exact endocytic pathway of VSV in the cell and on RABV transport in the neuronal axon. In any case, fusion is triggered in the endosomal vesicle via a low-pH induced structural rearrangement of G from its pre- to its postfusion conformation. Vesiculovirus G is one of the best characterized fusion glycoproteins as the previously reported crystal structures of the pre- and postfusion states have been recently completed by those of intermediates during the structural transition. Understanding the entry pathway of rhabdoviruses may have strong impact in biotechnologies as, for example, VSV G is used for pseudotyping lentiviruses to promote efficient transduction, and VSV is a promising oncolytic virus.
    Mots-clés : Chandipura virus, Clathrin-mediated endocytosis, Glycoprotein, LDL-receptor, Membrane fusion, Rabies virus, RHABDO, Rhabdovirus, Vesicular stomatitis virus, Viral receptor, VIRO.

  • L. Belot, A. Albertini, et Y. Gaudin, « Chapter Five - Structural and cellular biology of rhabdovirus entry », in Advances in Virus Research, vol. 104, M. Kielian, T. C. Mettenleiter, et M. J. Roossinck, Éd. Academic Press, 2019, p. 147-183.
    Résumé : Rhabdoviruses are enveloped viruses with a negative-sense single strand RNA genome and are widespread among a great variety of organisms. In their membrane, they have a single glycoprotein (G) that mediates both virus attachment to cellular receptors and fusion between viral and endosomal membranes allowing viral genome release in the cytoplasm. We present structural and cellular aspects of Rhabdovirus entry into their host cell with a focus on vesicular stomatitis virus (VSV) and rabies virus (RABV) for which the early events of the viral cycle have been extensively studied. Recent data have shown that the only VSV receptors are the members of the LDL-R family. This is in contrast with RABV for which multiple receptors belonging to unrelated families have been identified. Despite having different receptors, after attachment, rhabdovirus internalization occurs through clathrin-mediated endocytosis (CME) in an actin-dependent manner. There are still debates about the exact endocytic pathway of VSV in the cell and on RABV transport in the neuronal axon. In any case, fusion is triggered in the endosomal vesicle via a low-pH induced structural rearrangement of G from its pre- to its postfusion conformation. Vesiculovirus G is one of the best characterized fusion glycoproteins as the previously reported crystal structures of the pre- and postfusion states have been recently completed by those of intermediates during the structural transition. Understanding the entry pathway of rhabdoviruses may have strong impact in biotechnologies as, for example, VSV G is used for pseudotyping lentiviruses to promote efficient transduction, and VSV is a promising oncolytic virus.
    Mots-clés : Chandipura virus, Clathrin-mediated endocytosis, Glycoprotein, LDL-receptor, Membrane fusion, Rabies virus, RHABDO, Rhabdovirus, Vesicular stomatitis virus, Viral receptor, VIRO.

  • K. Ben Ouirane, Y. Boulard, et S. Bressanelli, « The hepatitis C virus RNA-dependent RNA polymerase directs incoming nucleotides to its active site through magnesium-dependent dynamics within its F motif », The Journal of Biological Chemistry, vol. 294, nᵒ 19, p. 7573-7587, mai 2019.
    Résumé : RNA viruses synthesize new genomes in the infected host thanks to dedicated, virally-encoded RNA-dependent RNA polymerases (RdRps). As such, these enzymes are prime targets for antiviral therapy, as has recently been demonstrated for hepatitis C virus (HCV). However, peculiarities in the architecture and dynamics of RdRps raise fundamental questions about access to their active site during RNA polymerization. Here, we used molecular modelling and molecular dynamics simulations, starting from the available crystal structures of HCV NS5B in ternary complex with template-primer duplexes and nucleotides, to address the question of ribonucleotide entry into the active site of viral RdRp. Tracing the possible passage of incoming UTP or GTP through the RdRp-specific entry tunnel, we found two successive checkpoints that regulate nucleotide traffic to the active site. We observed that a magnesium-bound nucleotide first binds next to the tunnel entry, and interactions with the triphosphate moiety orient it such that its base moiety enters first. Dynamics of the RdRp motifs F1 + F3 then allow the nucleotide to interrogate the RNA template base prior to nucleotide insertion into the active site. These dynamics are finely regulated by a second magnesium dication, thus coordinating the entry of a magnesium-bound nucleotide with shuttling of the second magnesium necessary for the two-metal ion catalysis. The findings of our work suggest that some at least of these features are general to viral RdRps and provide further details on the original nucleotide selection mechanism operating in RdRps of RNA viruses.
    Mots-clés : B3S, catalysis, complex, crystal-structure, fidelity, IMAPP, insights, mechanism, molecular dynamics, nucleoside, nucleoside/nucleotide transport, nucleotide transport, positive-sense RNA virus, protein motif, RNA virus, RNA-dependent RNA polymerase (RdRp), simulations, Single-stranded, Single-stranded, positive-sense RNA virus, structural basis, structural biology, viral polymerase.

  • L. Benkaidali, F. André, G. Moroy, B. Tangour, F. Maurel, et M. Petitjean, « Four Major Channels Detected in the Cytochrome P450 3A4: A Step toward Understanding Its Multispecificity », International Journal of Molecular Sciences, vol. 20, nᵒ 4, p. 987, févr. 2019.
    Résumé : We computed the network of channels of the 3A4 isoform of the cytochrome P450 (CYP) on the basis of 16 crystal structures extracted from the Protein Data Bank (PDB). The calculations were performed with version 2 of the CCCPP software that we developed for this research project. We identified the minimal cost paths (MCPs) output by CCCPP as probable ways to access to the buried active site. The algorithm of calculation of the MCPs is presented in this paper, with its original method of visualization of the channels. We found that these MCPs constitute four major channels in CYP3A4. Among the many channels proposed by Cojocaru et al. in 2007, we found that only four of them open in 3A4. We provide a refined description of these channels together with associated quantitative data.
    Mots-clés : access channels, active site access channels, B3S, buried active-site, cavities boundaries, cavities boundaries, crystal-structure, CYP3A4, LSOD.

  • E. Bordet, M. Frétaud, E. Crisci, E. Bouguyon, S. Rault, J. Pezant, A. Pleau, P. Renson, E. Giuffra, T. Larcher, M. Bourge, O. Bourry, O. Boulesteix, C. Langevin, I. Schwartz-Cornil, et N. Bertho, « Macrophage-B Cell Interactions in the Inverted Porcine Lymph Node and Their Response to Porcine Reproductive and Respiratory Syndrome Virus », Frontiers in Immunology, vol. 10, p. 953, 2019.
    Résumé : Swine lymph nodes (LN) present an inverted structure compared to mouse and human, with the afferent lymph diffusing from the center to the periphery. This structure, also observed in close and distant species such as dolphins, hippopotamus, rhinoceros, and elephants, is poorly described, nor are the LN macrophage populations and their relationship with B cell follicles. B cell maturation occurs mainly in LN B cell follicles with the help of LN macrophage populations endowed with different antigen delivery capacities. We identified three macrophage populations that we localized in the inverted LN spatial organization. This allowed us to ascribe porcine LN MΦ to their murine counterparts: subcapsular sinus MΦ, medullary cord MΦ and medullary sinus MΦ. We identified the different intra and extrafollicular stages of LN B cells maturation and explored the interaction of MΦ, drained antigen and follicular B cells. The porcine reproductive and respiratory syndrome virus (PRRSV) is a major porcine pathogen that infects tissue macrophages (MΦ). PRRSV is persistent in the secondary lymphoid tissues and induces a delay in neutralizing antibodies appearance. We observed PRRSV interaction with two LN MΦ populations, of which one interacts closely with centroblasts. We observed BCL6 up-regulation in centroblast upon PRRSV infection, leading to new hypothesis on PRRSV inhibition of B cell maturation. This seminal study of porcine LN will permit fruitful comparison with murine and human LN for a better understanding of normal and inverted LN development and functioning.
    Mots-clés : antibodies, antigen, B cell, BCL6, CYTO, PF, PRRSV.

  • L. Bossi, M. Ratel, C. Laurent, P. Kerboriou, A. Camilli, E. Eveno, M. Boudvillain, et N. Figueroa-Bossi, « NusG prevents transcriptional invasion of H-NS-silenced genes », PLoS genetics, vol. 15, nᵒ 10, p. e1008425, oct. 2019.
    Résumé : Evolutionarily conserved NusG protein enhances bacterial RNA polymerase processivity but can also promote transcription termination by binding to, and stimulating the activity of, Rho factor. Rho terminates transcription upon anchoring to cytidine-rich motifs, the so-called Rho utilization sites (Rut) in nascent RNA. Both NusG and Rho have been implicated in the silencing of horizontally-acquired A/T-rich DNA by nucleoid structuring protein H-NS. However, the relative roles of the two proteins in H-NS-mediated gene silencing remain incompletely defined. In the present study, a Salmonella strain carrying the nusG gene under the control of an arabinose-inducible repressor was used to assess the genome-wide response to NusG depletion. Results from two complementary approaches, i) screening lacZ protein fusions generated by random transposition and ii) transcriptomic analysis, converged to show that loss of NusG causes massive upregulation of Salmonella pathogenicity islands (SPIs) and other H-NS-silenced loci. A similar, although not identical, SPI-upregulated profile was observed in a strain with a mutation in the rho gene, Rho K130Q. Surprisingly, Rho mutation Y80C, which affects Rho's primary RNA binding domain, had either no effect or made H-NS-mediated silencing of SPIs even tighter. Thus, while corroborating the notion that bound H-NS can trigger Rho-dependent transcription termination in vivo, these data suggest that H-NS-elicited termination occurs entirely through a NusG-dependent pathway and is less dependent on Rut site binding by Rho. We provide evidence that through Rho recruitment, and possibly through other still unidentified mechanisms, NusG prevents pervasive transcripts from elongating into H-NS-silenced regions. Failure to perform this function causes the feedforward activation of the entire Salmonella virulence program. These findings provide further insight into NusG/Rho contribution in H-NS-mediated gene silencing and underscore the importance of this contribution for the proper functioning of a global regulatory response in growing bacteria. The complete set of transcriptomic data is freely available for viewing through a user-friendly genome browser interface.
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  • C. Bou-Nader, P. Barraud, L. Pecqueur, J. Perez, C. Velours, W. Shepard, M. Fontecave, C. Tisne, et D. Hamdane, « Molecular basis for transfer RNA recognition by the double-stranded RNA-binding domain of human dihydrouridine synthase 2 », Nucleic Acids Research, vol. 47, nᵒ 6, p. 3117-3126, avr. 2019.
    Résumé : Double stranded RNA-binding domain (dsRBD) is a ubiquitous domain specialized in the recognition of double-stranded RNAs (dsRNAs). Present in many proteins and enzymes involved in various functional roles of RNA metabolism, including RNA splicing, editing, and transport, dsRBD generally binds to RNAs that lack complex structures. However, this belief has recently been challenged by the discovery of a dsRBD serving as a major tRNA binding module for human dihydrouridine synthase 2 (hDus2), a flavoenzyme that catalyzes synthesis of dihydrouridine within the complex elbow structure of tRNA. We here unveil the molecular mechanism by which hDus2 dsRBD recognizes a tRNA ligand. By solving the crystal structure of this dsRBD in complex with a dsRNA together with extensive characterizations of its interaction with tRNA using mutagenesis, NMR and SAXS, we establish that while hDus2 dsRBD retains a conventional dsRNA recognition capability, the presence of an N-terminal extension appended to the canonical domain provides additional residues for binding tRNA in a structure-specific mode of action. Our results support that this extension represents a feature by which the dsRBD specializes in tRNA biology and more broadly highlight the importance of structural appendages to canonical domains in promoting the emergence of functional diversity.
    Mots-clés : complex reveals, evolution, extended dsrbd, mechanism, motif, nmr, PF, PIM, proteins, structural insights, trna(3)(lys).

  • A. Boussac, « Temperature dependence of the high-spin S-2 to S-3 transition in Photosystem II: Mechanistic consequences », Biochimica Et Biophysica Acta-Bioenergetics, vol. 1860, nᵒ 6, p. 508-518, juin 2019.
    Résumé : The Mn4CaO5-cluster in Photosystem II advances through five oxidation states, S-0 to S-4, before water is oxidized and O-2 is generated. The S-2-state exhibits either a low-spin, S = 1/2 (S-2(LS)), or a high-spin state, S = 5/2 (S-2(HS)). Increasing the pH favors the S-2(HS) Sens configuration and mimics the formation of Tyr(z)center dot in the S-2(LS)-state at lower pH values (Boussac et al. Biochim. Biophys. Acta 1859 (2018) 342). Here, the temperature dependence of the S-2(HS) to S-3 transition was studied by EPR spectroscopy at pH 8.6. The present data strengthened the involvement of S2I Us as a transient state in the S(2)(LS)Tyr(z)center dot S(2)(HS)Tyr(z) -> S(3)Tyr(z) transition. Depending on the temperature, the S-2(HS) progresses to S-3 states exhibiting different EPR properties. One S-3-state with a S = 3 signal, supposed to have a structure with the water molecule normally inserted in S-2 to S-3 transition, can be formed at temperatures as low as 77 K. This suggests that this water molecule is already bound in the S-2(HS) state at pH 8.6. The nature of the EPR invisible S-3 state, formed down to 4.2 K from a S-2(HS) is state, and that of the EPR detectable S3 state formed down to 77 K are discussed. It is proposed that in the S-2(LS) to S-3 transition, at pH < 8.6, the proton release (Sugiura et al. Biochim. Biophys. Acta 1859 (2018) 1259), the S-2(LS) to S-2(HS) conversion and the binding of the water molecule are all triggered by the formation of Tyr(z)center dot.
    Mots-clés : active-site, B3S, bond formation, camn4o5 cluster, electron-paramagnetic-res, EPR, Mn(4)CaO(5) cluster, Mn4CaO5 cluster, Oxygen evolution, Photosystem II, PS2, Spin state.

  • B. Bouznif, I. Guefrachi, R. C. Rodríguez de la Vega, M. Hungria, M. Mars, B. Alunni, et J. A. Shykoff, « Phylogeography of the Bradyrhizobium spp. Associated With Peanut, Arachis hypogaea: Fellow Travelers or New Associations? », Frontiers in Microbiology, vol. 10, p. 2041, 2019.
    Résumé : Legume plants have colonized almost all terrestrial biotopes. Their ecological success is partly due to the selective advantage provided by their symbiotic association with nitrogen-fixing bacteria called rhizobia, which allow legumes to thrive on marginal lands and nitrogen depleted soils where non-symbiotic plants cannot grow. Additionally, their symbiotic capacities result in a high protein content in their aerial parts and seeds. This interesting nutritional value has led to the domestication and agricultural exploitation of several legumes grown for seeds and/or fodder for human and domestic animal consumption. Several cultivated legume species are thus grown far beyond their natural geographic range. Other legume species have become invasives, spreading into new habitats. The cultivation and establishment of legume species outside of their original range requires either that they are introduced or cultivated along with their original symbiotic partner or that they find an efficient symbiotic partner in their introduced habitat. The peanut, Arachis hypogaea, a native of South America, is now cultivated throughout the world. This species forms root nodules with Bradyrhizobium, but it is unclear whether these came with the seeds from their native range or were acquired locally. Here we propose to investigate the phylogeography of Bradyrhizobium spp. associated with a number of different wild and cultivated legume species from a range of geographical areas, including numerous strains isolated from peanut roots across the areas of peanut cultivation. This will allow us to address the question of whether introduced/cultivated peanuts associate with bacteria from their original geographic range, i.e., were introduced together with their original bacterial symbionts, or whether they acquired their current associations de novo from the bacterial community within the area of introduction. We will base the phylogenetic analysis on sequence data from both housekeeping and core genes and a symbiotic gene (nif). Differences between the phylogenetic signal of symbiotic and non-symbiotic genes could result from horizontal transfer of symbiosis capacity. Thus this study will also allow us to elucidate the processes by which this symbiotic association has evolved within this group of Bradyrhizobium spp.
    Mots-clés : horizontal gene transfer, horizontal gene transfer (HGT), Host range, Legume-Rhizobium association, MICROBIO, PBI, Symbiosis.
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  • G. Briassoulis, P. Briassoulis, M. Miliaraki, S. Ilia, M. Parlato, F. Philippart, A. Rouquette, V. Moucadel, J. - M. Cavaillon, B. Misset, et Combined Approach for The eArly diagnosis of INfection in sepsis (CAPTAIN) study group, « Biomarker cruises in sepsis: who is the CAPTAIN? Discussion on "Circulating biomarkers may be unable to detect infection at the early phase of sepsis in ICU patients: the CAPTAIN prospective multicenter cohort study" », Intensive Care Medicine, janv. 2019.

  • A. Briquet, R. Vong, J. - B. Roseau, E. Javelle, N. Cazes, F. Rivière, M. Aletti, M. - P. Otto, C. Ficko, S. Duron, M. Fabre, C. Pourcel, F. Simon, et C. Soler, « Clinical features of Mycobacterium canettii infection: a retrospective study of 20 cases among French soldiers and relatives », Clinical Infectious Diseases: An Official Publication of the Infectious Diseases Society of America, févr. 2019.
    Résumé : Background: Mycobacterium canettii forms part of the Mycobacterium tuberculosis complex. M. canettii infections are mainly described in the Horn of Africa. The permanent presence of French soldiers in Djibouti raises the question of the risk of being infected with M. canettii. Our study aims to describe M. canettii infections among French military or their families between 1998 and 2015. Methods: This retrospective study relied on 3 sources of data: the reference centre for mycobacteria in the Biology Department at Percy military hospital in Paris, the French Military Center for Epidemiology and Public Health, and the scientific literature. After an exhaustive census of the strains, we studied the epidemiological data on 20 cases among French soldiers and their families. Results: 20 cases of M. canettii infections are reported, including 5 unpublished cases. Adenitis predominates (n = 15), especially in the cervico facial area and among children; one case was observed one month after dental care in Djibouti. The pulmonary forms were less frequent (n = 6) and 3 atypical forms are described. All patients had stayed in Djibouti. Conclusions: Cases of M. canettii infection among the French military consisted mainly of adenitis; disseminated forms were possible with immunodeficiency. Their evolution under specific treatments were comparable to tuberculosis. The presumed origin of the infection seemed to be environmental, possibly a water reservoir, and not due to human-to-human contagion.
    Mots-clés : DBG, LGBMB, MICROBIO, SSFA.

  • E. M. S. Brito, V. M. Romero-Núñez, C. A. Caretta, P. Bertin, J. C. Valerdi-Negreros, R. Guyoneaud, et M. Goñi-Urriza, « The bacterial diversity on steam vents from Paricutín and Sapichu volcanoes », Extremophiles: Life Under Extreme Conditions, févr. 2019.
    Résumé : Vapor steam vents are prevailing structures on geothermal sites in which local geochemical conditions allow the development of extremophilic microorganisms. We describe the structure of the prokaryotic community able to grow on the walls and rocks of such microecosystems in two terrestrial Mexican volcanoes: Paricutín (PI and PII samples) and its satellite Sapichu (S sample). The investigated samples showed similar diversity indices, with few dominant OTUs (abundance > 1%): 21, 16 and 23, respectively for PI, PII and S. However, each steam vent showed a particular community profile: PI was dominated by photosynthetic bacteria (Cyanobacteria and Chloroflexia class), PII by Actinobacteria and Proteobacteria, and S by Ktedonobacteria class, Acidobacteria and Cyanobacteria phyla. Concerning the predicted metabolic potential, we found a dominance of cellular pathways, especially the ones for energy generation with metabolisms for sulfur respiration, nitrogen fixation, methanogenesis, carbon fixation, photosynthesis, and metals, among others. We suggest a different maturity stage for the three studied fumaroles, from the youngest (PI) to the oldest (S and PII), also influenced by the temperature and other geochemical parameters. Furthermore, four anaerobic strains were isolated, belonging to Clostridia class (Clostridium sphenoides, C. swellfunanium and Anaerocolumna cellulosilytica) and to Bacilli class (Paenibacillus azoreducens).
    Mots-clés : Anaerobic bacteria, DBG, Extreme environment, GST, Microbial biodiversity, Predictive metagenomics profiling, Volcanic fumaroles.

  • A. Cabrie, O. Guittet, R. Tomasini, P. Vincendeau, et M. Lepoivre, « Crosstalk between TAp73 and TGF-beta in fibroblast regulates iNOS expression and Nrf2-dependent gene transcription », Free Radical Biology and Medicine, vol. 134, p. 617-629, avr. 2019.
    Résumé : Inducible nitric oxide synthase (iNOS) activity produces anti-tumor and anti-microbial effects but also promotes carcinogenesis through mutagenic, immunosuppressive and pro-angiogenic mechanisms. The tumor suppressor p53 contributes to iNOS downregulation by repressing induction of the NOS2 gene encoding iNOS, thereby limiting NO-mediated DNA damages. This study focuses on the role of the p53 homologue TAp73 in the regulation of iNOS expression. Induction of iNOS by immunological stimuli was upregulated in immortalized MEFs from TAp73(-/-) mice, compared to TAp73(+/+) fibroblasts. This overexpression resulted both from increased levels of NOS2 transcripts, and from an increased stability of the protein. Limitation of iNOS expression by TAp73 in wild-type cells is alleviated by TGF-beta receptor I inhibitors, suggesting a cooperation between TAp73 and TGF-beta in suppression of iNOS expression. Accordingly, downregulation of iNOS expression by exogenous TGF-beta 1 was impaired in TAp73(-/-) fibroblasts. Increased NO production in these cells resulted in a stronger, NO-dependent induction of Nrf2 target genes, indicating that the Nrf2-dependent adaptive response to nitrosative stress in fibroblasts is proportional to iNOS activity. NO-dependent induction of two HIF-1 target genes was also stronger in TAp73-deficient cells. Finally, the antimicrobial action of NO against Trypanosoma musculi parasites was enhanced in TAp73(-/-) fibroblasts. Our data indicate that tumor suppressive TAp73 isoforms cooperate with TGF-beta to control iNOS expression, NO-dependent adaptive responses to stress, and pathogen proliferation.
    Mots-clés : activation, B3S, cells, growth, immunity, Inducible nitric oxide synthase, innate, LBMS, macrophages, Nitric oxide, nitric-oxide production, Nuclear factor erythroid 2-related factor 2, suppression, Transforming growth factor beta, Transforming growth factor beta, transforming growth-factor-beta-1, tumorigenesis.

  • P. I. Calzadilla, F. Muzzopappa, P. Setif, et D. Kirilovsky, « Different roles for ApcD and ApcF in Synechococcus elongatus and Synechocystis sp. PCC 6803 phycobilisomes », Biochimica Et Biophysica Acta-Bioenergetics, vol. 1860, nᵒ 6, p. 488-498, juin 2019.
    Résumé : The phycobilisome, the cyanobacterial light harvesting complex, is a huge phycobiliprotein containing extra membrane complex, formed by a core from which rods radiate. The phycobilisome has evolved to efficiently absorb sun energy and transfer it to the photosystems via the last energy acceptors of the phycobilisome, ApcD and ApcE. ApcF also affects energy transfer by interacting with ApcE. In this work we studied the role of ApcD and ApcF in energy transfer and state transitions in Synechococcus elongatus and Synechocystis PCC6803. Our results demonstrate that these proteins have different roles in both processes in the two strains. The lack of ApcD and ApcF inhibits state transitions in Synechocystis but not in S. elongatus. In addition, lack of ApcF decreases energy transfer to both photosystems only in Synechocystis, while the lack of ApcD alters energy transfer to photosystem I only in S. elongatus. Thus, conclusions based on results obtained in one cyanobacterial strain cannot be systematically transferred to other strains and the putative role(s) of phycobilisomes in state transitions need to be reconsidered.
    Mots-clés : anacystis-nidulans, B3S, chlamydomonas-reinhardtii, Cyanobacteria, Energy transfer, excitation-energy transfer, light, MROP, orange carotenoid protein, photosystem-ii fluorescence, Phycobilisome, porphyridium-cruentum, quenching mechanisms, red alga, State transition, state transitions.

  • P. I. Calzadilla, J. Zhan, P. Sétif, C. Lemaire, D. Solymosi, N. Battchikova, Q. Wang, et D. Kirilovsky, « The cytochrome b6f complex is not involved in cyanobacterial state transitions », The Plant Cell, vol. 31, nᵒ 4, p. 911-931, avr. 2019.
    Résumé : Photosynthetic organisms need to sense and respond to fluctuating environmental conditions to avoid the formation of dangerous reactive oxygen species. The excitation energy arriving at each photosystem permanently changes due to variations of intensity and spectral properties of the absorbed light. Cyanobacteria, like plants and algae, have developed a mechanism, named state transitions, that sense and respond to these fluctuating conditions. In this work, we characterize the role of the cytochrome b6f and phosphorylation reactions in cyanobacterial state transitions using Synechococcus elongatus PCC 7942 and Synechocystis PCC 6803. A large Photosystem II fluorescence quenching was observed in State II which seems not to be related to spillover. This membrane-associated process was inhibited by betaine, sucrose and high concentrations of phosphate. Then, using different chemicals affecting the PQ pool redox state and the activity of the cytochrome b6f, we demonstrated that this complex is not involved in S. elongatus and Synechocystis PCC6803 state transitions. Finally, by constructing and characterizing 21 kinase and phosphatase mutants and using chemical inhibitors, it was clearly shown that phosphorylation reactions are not essential in cyanobacterial state transitions. Thus, signal transduction is completely different in cyanobacteria and plant (green alga) state transitions.
    Mots-clés : B3S, MROP.

  • V. Campanacci, A. Urvoas, S. Cantos-Fernandes, M. Aumont-Nicaise, A. - A. Arteni, C. Velours, M. Valerio-Lepiniec, B. Dreier, A. Plückthun, A. Pilon, C. Poüs, P. Minard, et B. Gigant, « Insight into microtubule nucleation from tubulin-capping proteins », Proceedings of the National Academy of Sciences of the United States of America, vol. 116, nᵒ 20, p. 9859-9864, avr. 2019.
    Résumé : Nucleation is one of the least understood steps of microtubule dynamics. It is a kinetically unfavorable process that is templated in the cell by the γ-tubulin ring complex or by preexisting microtubules; it also occurs in vitro from pure tubulin. Here we study the nucleation inhibition potency of natural or artificial proteins in connection with their binding mode to the longitudinal surface of α- or β-tubulin. The structure of tubulin-bound CopN, a Chlamydia protein that delays nucleation, suggests that this protein may interfere with two protofilaments at the (+) end of a nucleus. Designed ankyrin repeat proteins that share a binding mode similar to that of CopN also impede nucleation, whereas those that target only one protofilament do not. In addition, an αRep protein predicted to target two protofilaments at the (-) end does not delay nucleation, pointing to different behaviors at both ends of the nucleus. Our results link the interference with protofilaments at the (+) end and the inhibition of nucleation.
    Mots-clés : artificial binding proteins, B3S, CopN, CRYOEM, MIKICA, MIP, PF, PIM.

  • V. Campanacci, A. Urvoas, T. Consolati, S. Cantos-Fernandes, M. Aumont-Nicaise, M. Valerio-Lepiniec, T. Surrey, P. Minard, et B. Gigant, « Selection and Characterization of Artificial Proteins Targeting the Tubulin alpha Subunit », Structure, vol. 27, nᵒ 3, p. 497-+, mars 2019.
    Résumé : Microtubules are cytoskeletal filaments of eukaryotic cells made of alpha beta-tubulin heterodimers. Structural studies of non-microtubular tubulin rely mainly on molecules that prevent its self-assembly and are used as crystallization chaperones. Here we identified artificial proteins from an alpha Rep library that are specific to alpha-tubulin. Turbidity experiments indicate that these alpha Reps impede microtubule assembly in a dose-dependent manner and total internal reflection fluorescence microscopy further shows that they specifically block growth at the microtubule (-) end. Structural data indicate that they do so by targeting the alpha-tubulin longitudinal surface. Interestingly, in one of the complexes studied, the alpha subunit is in a conformation that is intermediate between the ones most commonly observed in X-ray structures of tubulin and those seen in the microtubule, emphasizing the plasticity of tubulin. These alpha-tubulin-specific alpha Reps broaden the range of tools available for the mechanistic study of microtubule dynamics and its regulation.
    Mots-clés : artificial protein, B3S, beta-tubulin, complex, depolymerization, design, dynamic instability, in vitro selection, microtubule, microtubule plus, MIKICA, MIP, overexpression, purification, stathmin, structural basis, tubulin, αRep.

  • N. Canu, M. Moutiez, P. Belin, et M. Gondry, « Cyclodipeptide synthases: a promising biotechnological tool for the synthesis of diverse 2,5-diketopiperazines », Natural Product Reports, août 2019.
    Résumé : Covering: Up to mid-2019Cyclodipeptide synthases (CDPSs) catalyse the formation of cyclodipeptides using aminoacylated-tRNA as substrates. The recent characterization of large sets of CDPSs has revealed that they can produce highly diverse products, and therefore have great potential for use in the production of different 2,5-diketopiperazines (2,5-DKPs). Sequence similarity networks (SSNs) are presented as a new, efficient way of classifying CDPSs by specificity and identifying new CDPS likely to display novel specificities. Several strategies for further increasing the diversity accessible with these enzymes are discussed here, including the incorporation of non-canonical amino acids by CDPSs and use of the remarkable diversity of 2,5-DKP-tailoring enzymes discovered in recent years.
    Mots-clés : BIOSYN, MICROBIO.

  • C. Carvalho, V. L'Hôte, R. Courbeyrette, G. Kratassiouk, G. Pinna, J. - C. Cintrat, C. Denby-Wilkes, C. Derbois, R. Olaso, J. - F. Deleuze, C. Mann, et J. - Y. Thuret, « Glucocorticoids delay RAF-induced senescence promoted by EGR1 », Journal of Cell Science, vol. 132, nᵒ 16, p. UNSP jcs230748, août 2019.
    Résumé : Expression of hyperactive RAF kinases, such as the oncogenic B-RAF-V600E mutant, in normal human cells triggers a proliferative arrest that blocks tumor formation. We discovered that glucocorticoids delayed the entry into senescence induced by B-RAF-V600E in human fibroblasts, and allowed senescence bypass when the cells were regularly passaged, but that they did not allow proliferation of cells that were already senescent. Transcriptome and siRNA analyses revealed that the EGR1 gene is one target of glucocorticoid action. Transcription of the EGR1 gene is activated by the RAF-MEK-ERK MAPK pathway and acts as a sensor of hyper-mitogenic pathway activity. The EGR1 transcription factor regulates the expression of p15 and p21 (encoded by CDKN2B and CDKN1A, respectively) that are redundantly required for the proliferative arrest of BJ fibroblasts upon expression of B-RAF-V600E. Our results highlight the need to evaluate the action of glucocorticoid on cancer progression in melanoma, thyroid and colon carcinoma in which B-RAF-V600E is a frequent oncogene, and cancers in which evasion from senescence has been shown.
    Mots-clés : B-RAF-V600E, CDKN1A, CDKN2B, DBG, EGR1, Glucocorticoid, GTR, Oncogene-induced senescence, PARI, PF, SEN.

  • R. Catchpole et P. Forterre, « The evolution of Reverse Gyrase suggests a non-hyperthermophilic Last Universal Common Ancestor », Molecular Biology and Evolution, sept. 2019.
    Résumé : Reverse gyrase (RG) is the only protein found ubiquitously in hyperthermophilic organisms, but absent from mesophiles. As such, its simple presence or absence allows us to deduce information about the optimal growth temperature of long-extinct organisms, even as far as the last universal common ancestor of extant life (LUCA). The growth environment and gene content of the LUCA has long been a source of debate in which RG often features. In an attempt to settle this debate, we carried out an exhaustive search for RG proteins, generating the largest RG dataset to date. Comprising 376 sequences, our dataset allows for phylogenetic reconstructions of RG with unprecedented size and detail. These RG phylogenies are strikingly different from those of universal proteins inferred to be present in the LUCA, even when using the same set of species. Unlike such proteins, RG does not form monophyletic archaeal and bacterial clades, suggesting RG emergence after the formation of these domains, and/or significant horizontal gene transfer. Additionally, the branch lengths separating archaeal and bacterial groups are very short, inconsistent with the tempo of evolution from the time of the LUCA. Despite this, phylogenies limited to archaeal RG resolve most archaeal phyla, suggesting predominantly vertical evolution since the time of the last archaeal ancestor. In contrast, bacterial RG indicates emergence after the last bacterial ancestor followed by significant horizontal transfer. Taken together, these results suggest a non-hyperthermophilic LUCA and bacterial ancestor, with hyperthermophily emerging early in the evolution of the archaeal and bacterial domains.
    Mots-clés : ARCHEE, MICROBIO.

  • P. Champeil, B. de Foresta, M. Picard, C. Gauron, D. Georgin, M. le Maire, J. V. Møller, G. Lenoir, et C. Montigny, « Interaction of detergents with biological membranes: Comparison of fluorescence assays with filtration protocols and implications for the rates of detergent association, dissociation and flip-flop », PloS One, vol. 14, nᵒ 10, p. e0222932, 2019.
    Résumé : The present study mainly consists of a re-evaluation of the rate at which C12E8, a typical non-ionic detergent used for membrane studies, is able to dissociate from biological membranes, with sarcoplasmic reticulum membrane vesicles being used as an example. Utilizing a brominated derivative of C12E8 and now stopped-flow fluorescence instead of rapid filtration, we found that the rate of dissociation of this detergent from these membranes, merely perturbed with non-solubilizing concentrations of detergent, was significantly faster (t1/2 < 10 ms) than what had previously been determined (t1/2 ~300-400 ms) from experiments based on a rapid filtration protocol using 14C-labeled C12E8 and glass fiber filters (Binding of a non-ionic detergent to membranes: flip-flop rate and location on the bilayer, by Marc le Maire, Jesper Møller and Philippe Champeil, Biochemistry (1987) Vol 26, pages 4803-4810). We here pinpoint a methodological problem of the earlier rapid filtration experiments, and we suggest that the true overall dissociation rate of C12E8 is indeed much faster than previously thought. We also exemplify the case of brominated dodecyl-maltoside, whose kinetics for overall binding to and dissociation from membranes comprise both a rapid and a sower phase, the latter being presumably due to flip-flop between the two leaflets of the membrane. Consequently, equilibrium is reached only after a few seconds for DDM. This work thereby emphasizes the interest of using the fluorescence quenching associated with brominated detergents for studying the kinetics of detergent/membrane interactions, namely association, dissociation and flip-flop rates.
    Mots-clés : B3S, LPSM.
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  • M. Chan-Yao-Chong, C. Deville, L. Pinet, C. van Heijenoort, D. Durand, et T. Ha-Duong, « Structural Characterization of N-WASP Domain V Using MD Simulations with NMR and SAXS Data », Biophysical Journal, vol. 116, nᵒ 7, p. 1216-1227, avr. 2019.
    Résumé : Because of their large conformational heterogeneity, structural characterization of intrinsically disordered proteins (IDPs) is very challenging using classical experimental methods alone. In this study, we use NMR and small-angle x-ray scattering (SAXS) data with multiple molecular dynamics (MD) simulations to describe the conformational ensemble of the fully disordered verprolin homology domain of the neural Aldrich syndrome protein involved in the regulation of actin polymerization. First, we studied several back-calculation software of SAXS scattering intensity and optimized the adjustable parameters to accurately calculate the SAXS intensity from an atomic structure. We also identified the most appropriate force fields for MD simulations of this IDP. Then, we analyzed four conformational ensembles of neural Aldrich syndrome protein verprolin homology domain, two generated with the program flexible-meccano with or without NMR-derived information as input and two others generated by MD simulations with two different force fields. These four conformational ensembles were compared to available NMR and SAXS data for validation. We found that MD simulations with the AMBER-03w force field and the TIP4P/2005s water model are able to correctly describe the conformational ensemble of this 67-residue IDP at both local and global level.
    Mots-clés : alpha-synuclein, angle scattering data, atomic-resolution, B3S, c-13' chemical-shifts, FAAM, force-field, fuzzy complexes, intrinsically disordered proteins, molecular recognition features, quantum-mechanics, unstructured proteins.

  • M. Chan-Yao-Chong, D. Durand, et T. Ha-Duong, « Molecular Dynamics Simulations Combined with Nuclear Magnetic Resonance and/or Small-Angle X-ray Scattering Data for Characterizing Intrinsically Disordered Protein Conformational Ensembles », Journal of Chemical Information and Modeling, vol. 59, nᵒ 5, p. 1743-1758, mai 2019.
    Résumé : The concept of intrinsically disordered proteins (IDPs) has emerged relatively slowly, but over the past 20 years, it has become an intense research area in structural biology. Indeed, because of their considerable flexibility and structural heterogeneity, the determination of IDP conformational ensemble is particularly challenging and often requires a combination of experimental measurements and computational approaches. With the improved accuracy of all-atom force fields and the increasing computing performances, molecular dynamics (MD) simulations have become more and more reliable to generate realistic conformational ensembles. And the combination of MD simulations with experimental approaches, such as nuclear magnetic resonance (NMR) and/or small-angle X-ray scattering (SAXS) allows one to converge toward a more accurate and exhaustive description of IDP structures. In this Review, we discuss the state of the art of MD simulations of IDP conformational ensembles, with a special focus on studies that back-calculated and directly compared theoretical and experimental NMR or SAXS observables, such as chemical shifts (CS), 3J-couplings (3Jc), residual dipolar couplings (RDC), or SAXS intensities. We organize the review in three parts. In the first section, we discuss the studies which used NMR and/or SAXS data to test and validate the development of force fields or enhanced sampling techniques. In the second part, we explore different methods for the refinement of MD-derived structural ensembles, such as NMR or SAXS data-restrained MD simulations or ensemble reweighting to better fit experiments. Finally, we survey some recent studies combining MD simulations with NMR and/or SAXS measurements to investigate the relationship between IDP conformational ensemble and biological activity, as well as their implication in human diseases. From this review, we noticed that quite a few studies compared MD-generated conformational ensembles with both NMR and SAXS measurements to validate IDP structures at both local and global levels. Yet, beside the IDP propensity to form local secondary structures, their dynamic extension or compactness also appears important for their activity. Thus, we believe that a close synergy between MD simulations, NMR, and SAXS experiments would be greatly appropriate to address the challenges of characterizing the disordered structures of proteins and their complexes, relative to their biological functions.
    Mots-clés : alpha-synuclein, atomic-level characterization, B3S, chemical-shifts, FAAM, folding simulations, force-field, generalized born model, replica exchange, structural ensembles, unfolded states, water model.

  • M. Chan-Yao-Chong, D. Durand, et T. Ha-Duong, « Investigation into Early Steps of Actin Recognition by the Intrinsically Disordered N-WASP Domain V », International Journal of Molecular Sciences, vol. 20, nᵒ 18, sept. 2019.
    Résumé : Cellular regulation or signaling processes are mediated by many proteins which often have one or several intrinsically disordered regions (IDRs). These IDRs generally serve as binders to different proteins with high specificity. In many cases, IDRs undergo a disorder-to-order transition upon binding, following a mechanism between two possible pathways, the induced fit or the conformational selection. Since these mechanisms contribute differently to the kinetics of IDR associations, it is important to investigate them in order to gain insight into the physical factors that determine the biomolecular recognition process. The verprolin homology domain (V) of the Neural Wiskott-Aldrich Syndrome Protein (N-WASP), involved in the regulation of actin polymerization, is a typical example of IDR. It is composed of two WH2 motifs, each being able to bind one actin molecule. In this study, we investigated the early steps of the recognition process of actin by the WH2 motifs of N-WASP domain V. Using docking calculations and molecular dynamics simulations, our study shows that actin is first recognized by the N-WASP domain V regions which have the highest propensity to form transient α -helices. The WH2 motif consensus sequences "LKKV" subsequently bind to actin through large conformational changes of the disordered domain V.
    Mots-clés : B3S, FAAM, intrinsically disordered protein, molecular docking, molecular dynamics, protein–protein interaction.
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  • F. Chauffour, M. Bailly, F. Perreau, G. Cueff, H. Suzuki, B. Collet, A. Frey, G. Clément, L. Soubigou-Taconnat, T. Balliau, A. Krieger-Liszkay, L. Rajjou, et A. Marion-Poll, « Multi-omics analysis reveals sequential roles for ABA during seed maturation », Plant Physiology, vol. 180, nᵒ 2, p. 1198-1218, avr. 2019.
    Résumé : Abscisic acid (ABA) is an important hormone for seed development and germination whose physiological action is modulated by its endogenous levels. Cleavage of carotenoid precursors by 9-cis epoxycarotenoid dioxygenase (NCED) and inactivation of ABA by ABA 8'-hydroxylase (CYP707A) are key regulatory metabolic steps. In Arabidopsis (Arabidopsis thaliana), both enzymes are encoded by multigene families, having distinctive expression patterns. To evaluate the genome-wide impact of ABA deficiency in developing seeds at the maturation stage when dormancy is induced, we used a nced2569 quadruple mutant in which ABA deficiency is mostly restricted to seeds, thus limiting the impact of maternal defects on seed physiology. ABA content was very low in nced2569 seeds, similar to the severe mutant aba2; unexpectedly, ABA glucose ester was detected in aba2 seeds, suggesting the existence of an alternative metabolic route. Hormone content in nced2569 seeds compared with nced259 and wild-type strongly suggested that specific expression of NCED6 in the endosperm is mainly responsible for ABA production. In accordance, transcriptome analyses revealed broad similarities in gene expression between nced2569 and either wild type or nced259 developing seeds. Gene ontology enrichments revealed a large spectrum of ABA activation targets involved in reserve storage and desiccation tolerance, and repression of photosynthesis and cell cycle. Proteome and metabolome profiles in dry nced2569 seeds, compared with wild-type and cyp707a1a2 seeds, also highlighted an inhibitory role of ABA on remobilisation of reserves, ROS production, and protein oxidation. Down-regulation of these oxidative processes by ABA may have an essential role in dormancy control.
    Mots-clés : 9-cis-epoxycarotenoid dioxygenase, abscisic-acid biosynthesis, arabidopsis seeds, B3S, dormancy, drought tolerance, genome-wide analysis, mass-spectrometry, metabolism, MROP, protein oxidation, signaling networks.

  • C. Chen, B. Corry, L. Huang, et N. Hildebrandt, « FRET-Modulated Multihybrid Nanoparticles for Brightness-Equalized Single-Wavelength Barcoding », Journal of the American Chemical Society, vol. 141, nᵒ 28, p. 11123-11141, juin 2019.
    Résumé : Semiconductor quantum dots (QDs) are the most versatile fluorophores for Förster resonance energy transfer (FRET) because they can function as both donors and acceptors for a multitude of fluorophores. However, a complete understanding of multidonor-multiacceptor FRET networks on QDs and their full employment into advanced fluorescence sensing and imaging have not been accomplished. Here, we provide a holistic photophysical analysis of such multidonor-QD-multiacceptor FRET systems using time-resolved and steady-state photoluminescence (PL) spectroscopy and Monte Carlo simulations. Multiple terbium complex (Tb) donors (1-191 units) and Cy5.5 dye acceptors (1-60 units) were attached to a central QD, and the entire range of combinations of FRET pathways was investigated by Tb, QD, and Cy5.5 PL. Experimental and simulation results were in excellent agreement and could disentangle the distinct contributions of hetero-FRET, homo-FRET, and dye dimerization. The FRET efficiency was independent of the number of Tb donors and dependent on the number of Cy5.5 acceptors, which could be used to independently adapt the PL intensity by the number of Tb donors and the PL lifetime by the number of Cy5.5 acceptors. We used this unique tuning capability to prepare Tb-QD-Cy5.5 conjugates with distinct QD PL lifetimes but similar QD PL intensities. These brightness-equalized multihybrid FRET nanoparticles were applied to optical barcoding via three time-gated PL intensity detection windows, which resulted in simple RGB ratios. Direct applicability was demonstrated by an efficient RGB distinction of different nanoparticle-encoded microbeads within the same field of view with both single-wavelength excitation and detection on a standard fluorescence microscope.
    Mots-clés : B3S, NANO.

  • J. - H. Chen, L. - J. Yu, A. Boussac, Z. - Y. Wang-Otomo, T. Kuang, et J. - R. Shen, « Properties and structure of a low-potential, penta-heme cytochrome c(552) from a thermophilic purple sulfur photosynthetic bacterium Thermochromatium tepidum », Photosynthesis Research, vol. 139, nᵒ 1-3, p. 281-293, mars 2019.
    Résumé : The thermophilic purple sulfur bacterium Thermochromatium tepidum possesses four main water-soluble redox proteins involved in the electron transfer behavior. Crystal structures have been reported for three of them: a high potential iron-sulfur protein, cytochrome c, and one of two low-potential cytochrome c(552) (which is a flavocytochrome c) have been determined. In this study, we purified another low-potential cytochrome c(552) (LPC), determined its N-terminal amino acid sequence and the whole gene sequence, characterized it with absorption and electron paramagnetic spectroscopy, and solved its high-resolution crystal structure. This novel cytochrome was found to contain five c-type hemes. The overall fold of LPC consists of two distinct domains, one is the five heme-containing domain and the other one is an Ig-like domain. This provides a representative example for the structures of multiheme cytochromes containing an odd number of hemes, although the structures of multiheme cytochromes with an even number of hemes are frequently seen in the PDB database. Comparison of the sequence and structure of LPC with other proteins in the databases revealed several characteristic features which may be important for its functioning. Based on the results obtained, we discuss the possible intracellular function of this LPC in Tch. tepidum.
    Mots-clés : angstrom resolution, B3S, c nitrite reductase, c554, conservation, Crystal structure, crystal-structure, Cytochrome c, Electron transfer, environment, genes, Multiheme, proteins, PS2, Purple sulfur bacteria, spectroscopy, subunit, Thermochromatium tepidum.

  • P. Chervy, C. Petcut, D. Rault, C. Meriadec, T. Bizien, K. François, J. Richard, C. Chassaing, N. Benamar, F. Artzner, et M. Paternostre, « Organic Nanoscrolls from Electrostatic Interactions between Peptides and Lipids: Assembly Steps and Structure », Langmuir, vol. 35, nᵒ 32, p. 10648-10657, août 2019.
    Résumé : An important aspect of cells is their shape flexibility that gives them motion but also a high adaptation versatility to their environment. This shape versatility is mediated by different types of protein–membrane interactions among which electrostatic plays an important role. In the present work we examined the interaction between a small dicationic peptide, that possesses self-assembly properties, and lipid model membranes. The peptide, lanreotide, spontaneously forms nanotubes in water that have a strictly uniform diameter. In the current work, we show that the interaction between the cationic peptide and negatively charged bilayers of lipids induces the formation of myelin sheath-like structures that we call nanoscrolls. By deciphering the different steps of formation and the molecular structure of the self-assembly, we show how electrostatics modify the spontaneous peptide and lipid way of packing.
    Mots-clés : B3S, IMAPP.

  • D. Ciardo, A. Goldar, et K. Marheineke, « On the Interplay of the DNA Replication Program and the Intra-S Phase Checkpoint Pathway », Genes, vol. 10, nᵒ 2, p. 94, janv. 2019.
    Résumé : DNA replication in eukaryotes is achieved by the activation of multiple replication origins which needs to be precisely coordinated in space and time. This spatio-temporal replication program is regulated by many factors to maintain genome stability, which is frequently threatened through stresses of exogenous or endogenous origin. Intra-S phase checkpoints monitor the integrity of DNA synthesis and are activated when replication forks are stalled. Their activation leads to the stabilization of forks, to the delay of the replication program by the inhibition of late firing origins, and the delay of G2/M phase entry. In some cell cycles during early development these mechanisms are less efficient in order to allow rapid cell divisions. In this article, we will review our current knowledge of how the intra-S phase checkpoint regulates the replication program in budding yeast and metazoan models, including early embryos with rapid S phases. We sum up current models on how the checkpoint can inhibit origin firing in some genomic regions, but allow dormant origin activation in other regions. Finally, we discuss how numerical and theoretical models can be used to connect the multiple different actors into a global process and to extract general rules.
    Mots-clés : ataxia-telangiectasia, ATR, Chk1, DBG, DNA replication, DYNREP, GTR, MBT.

  • E. C. Conceição, G. Refregier, H. M. Gomes, X. Olessa-Daragon, F. Coll, N. H. Ratovonirina, V. Rasolofo-Razanamparany, M. L. Lopes, D. van Soolingen, L. Rutaihwa, S. Gagneux, V. R. Bollela, P. N. Suffys, R. S. Duarte, K. V. B. Lima, et C. N. Sola, « Mycobacterium tuberculosis lineage 1 genetic diversity in Pará, Brazil, suggests common ancestry with east-African isolates potentially linked to historical slave trade », Infection, Genetics and Evolution: Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases, vol. 73, p. 337-341, juin 2019.
    Résumé : Lineage 1 (L1) is one of seven Mycobacterium tuberculosis complex (MTBC) lineages. The objective of this study was to improve the complex taxonomy of L1 using phylogenetic SNPs, and to look for the origin of the main L1 sublineage prevalent in Para, Brazil. We developed a high-throughput SNPs-typing assay based on 12-L1-specific SNPs. This assay allowed us to experimentally retrieve SNP patterns on nine of these twelve SNPs in 277 isolates previously tentatively assigned to L1 spoligotyping-based sub lineages. Three collections were used: Pará-Brazil (71); RIVM, the Netherlands (102), Madagascar (104). One-hundred more results were generated in Silico using the PolyTB database. Based on the final SNPs combination, the samples were classified into 11 clusters (C1-C11). Most isolates within a SNP-based cluster shared a mutual spoligotyping-defined lineage. However, L1/EAI1-SOM (SIT48, sp. 40) and L1/EAI6-BGD1 (SIT591, sp. 23) showed a poor correlation with SNP data and are not monophyletic. L1/EAI8-MDG and L1/EAI3-IND belonged to C5; this result suggests that they share a common ancestor. L1.1.3/SIT129, a spoligotype pattern found in SNPs-cluster C6, was found to be shared between Pará/Brazil and Malawi. SIT129 was independently found to be highly prevalent in Mozambique, which suggests a migration history from East-Africa to Brazil during the 16th-18th slave trade period to Northern Brazil.
    Mots-clés : IGEPE, Lineage 1, MICROBIO, Molecular evolution, Mycobacterium tuberculosis complex, Single-nucleotide polymorphisms, Spoligotyping, Whole genome sequencing.

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