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Legumes provide several ecological services to agroecosystems, but there is a lack of references on services related to N flows for a wide range of legume crops. N2O emissions were measured in two field experiments using a two-year legume-cereal crop sequence. In the first year (2014 and 2016), different legume crops were grown (lupin, pea, fava bean, common bean, soybean, chickpea) and compared to fertilized cereals (barley and sorghum). Once the seeds were harvested and the residues incorporated in the soil, unfertilized wheat was sown and harvested in the second year (2015 and 2017). N2O emissions, as well as soil temperature and moisture, were measured continuously using an automated chamber method during the two years of each experiment. Daily N2O emissions were less than 10 g N-N2O ha-1 d-1, with higher values (ranging from 10 to 90 g N-N2O ha-1 d-1) being measured during exceptionally rainy conditions. Daily N2O emissions were mainly influenced by climatic conditions for field experiments and far less by inorganic N content, except for N-fertilized cereals. For both field experiments, cumulative N2O emissions during legume and cereal pre-crops + fallow period between pre-crop harvest and wheat sowing (1st year) (mean values 365.4 and 318.1 g N-N2O ha-1 for experiment I and II, respectively) were higher than during wheat crop cultivation (2nd year) (155.8 and 101.5 g N-N2O ha-1 for experiment I and II, respectively). For field experiment II, N2O emissions were slightly higher for the N fertilized cereal pre-crops (529.8 and 523.3 g N-N2O ha-1 for barley and sorghum, respectively) compared to legume pre-crops (mean values 380.6 and 417.2 g N-N2O ha-1 for legumes sown in March and May, respectively), while no significant difference was measured for field experiment I. There was no difference in N2O emissions during the cultivation of the different legume species. Furthermore, when wheat was grown after legumes or N fertilized cereals, N2O emissions were comparable for the different experimental treatments with no relation established with the amounts of N present in crop residues or their C: N ratios. Despite the small differences in emissions between N-fertilized cereals and grain legumes, introducing these leguminous species in crop rotation and in these pedoclimatic conditions makes it possible to substitute synthetic N fertilizer and mitigate the greenhouse gases emitted from these cropping systems. However, further research is still needed to clarify and quantify the value of legumes in mitigating and reducing greenhouse gas emissions from cropping systems.

International audience; The development of inhibitors of anti-apoptotic proteins, such as Mcl-1, is currently a very active area in the field of cancer research. One of the very first reported inhibitor of Mcl-1 was the molecule MIM1, but we have demonstrated recently that the structure of this compound had to be revised from 2 to the derivative 1 (FJ-809). In this paper we first develop a strategy to prepare unambiguously molecules such as 1 with a thiazol-3(2H)-yl)imino core, instead of the [2(3H)thiazolylidene]hydrazine previously found in MIM1 (2). Next a series of biological studies have been performed on 1, using IGROV1-R10 ovarian cancer cells as models, and they have been complemented by Fluorescence Polarisation Assays. These studies demonstrated that the new compound FJ-809 (1) was devoid of any significant activity on Mcl-1, contrary to 2. Then molecular modelling and molecular dynamic studies have been performed in order to elucidate the differences between FJ-809 and MIM1 in their interaction with the Mcl-1 protein.

International audience; In the area of cancer research, the development of new and potent inhibitors of anti-apoptotic proteins is a very active and promising topic. The small molecule MIM1 has been reported earlier as one of the first selective inhibitors of the anti-apoptotic protein Mcl-1. In the present paper, we first revised the structure of this molecule based on extensive physicochemical analyses. Then we designed and synthesized a focused library of analogues for the corrected structure of MIM1. Next, these molecules were subjected to a panel of in cellulo biological studies, allowing the identification of dual Bcl-x(L)/Mcl-1 inhibitors, as well as selective Mcl-1 inhibitors. These results have been complemented by fluorescence polarization assays with the Mcl-1 protein. Preliminary structure-activity relationships were discussed and extensive molecular modelling studies allowed us to propose a rationale for the biological activity of this series of new inhibitors, in particular for the selectivity of inhibition of Mcl-1 versus Bcl-x(L).

In this study, we explored the structural dynamics of Mcl-1, an anti-apoptotic protein. On the basis of structural ensembles, the essential dynamics was extracted and showed two major axes of variability: a breathing motion at the binding interface and a correlated motion through the internal loops. A free energy surface characterizing the breathing motion at the binding interface was generated and suggested an equilibrium between a closed conformation and a "ready to bind" conformation as the predominant states of Mcl-1 in solution. Moreover, the analysis of the dynamics along the internal loops revealed a hidden communication network of transient and cryptic pockets controlling the allosteric inhibition of Mcl-1. A detailed model joining the pocket crosstalk and salt bridge networks along the internal loops was proposed and allowed us to shed light on the key interactions governing Mcl-1's allosteric inhibition.

Apoptosis is a highly regulated cellular process. Aberration in apoptosis is a common characteristic of various disorders. Therefore, proteins involved in apoptosis are prime targets in multiple therapies. Bcl-x

Detection of cryptic pockets (hidden protein pockets) is a hot topic in structure-based drug discovery, especially for drugging the yet undruggable proteome. The experimental detection of cryptic pockets is still considered an expensive endeavor. Thus, computational methods, such as atomistic simulations, are used instead. These simulation methods can provide a perspective on protein dynamics that overpasses the experimental X-ray structures' static and average view. Nonetheless, unbiased molecular dynamics (MD) simulations fall short to detect transient and cryptic pockets requiring the crossing of high-energy barriers. Enhanced sampling methods, such as Metadynamics, provide a solution to overcome the time-scale problem faced by unbiased MD simulations. However, these methods are still limited by the availability of collective variable space to capture the intricate parameters, leading to the opening of cryptic pockets. Unfortunately, the design of such collective variables requires a priori knowledge of the binding site, information that is by definition lacking for cryptic pockets. In this work, we evaluated the use of the Metadynamics biasing scheme on essential coordinates space as a general method for cryptic pocket detection. This approach was applied to an antiapoptotic protein: Mcl-1 as a test model. In addition to providing a broader characterization of Mcl-1's conformational space, we show the effectiveness of this method in drawing the full repository of Mcl-1's known and novel cryptic pockets in an unsupervised manner.

Target Protein Degradation TPD is a new avenue and revolutionary for therapeutics because redefining the principles of classical drug discovery and guided by event-based target activity rather than the occupancy-driven activity. Since the discovery of the first PROTAC in 2001, TPD represents a rapidly growing technology, with applications in both drug discovery and chemical biology. Over the last decade, many questions have been raised and today the knowledge gained by each team has elucidated a number of them, although there is still a long way to go. The objective of this work is to present the challenges that the PROTAC strategy has very recently addressed in drug design and discovery by presenting extremely recent results from the literature and to provide guidelines in the drug design of new PROTACs as successful therapeutic modality for medicinal chemists.

Tauopathies are neurodegenerative disorders associated with the accumulation of abnormal tubulin associated unit (tau) protein in the brain. Tau pathologies include a broad spectrum of diseases, with Alzheimer's disease (AD) being the most common tauopathy. The pathophysiological mechanisms of AD are still only partially understood. As a consequence, attempts to establish therapeutic approaches have led to numerous clinical trial failures and, to date, no curative treatment is available for AD despite the considerable number of research programs. Therefore, over the past decade, the aggregation of the tau protein in AD has become a therapeutic target of interest. In this review, we gather in silico, in vitro, and in vivo methodologies that are relevant to assess compounds targeting tau aggregation, from early drug design to clinical trials.

Protein-protein interactions (PPIs) control many important physiological processes within human cells. Apoptosis or programmed cell death is closely regulated by pro- and antiapoptotic signals. Dysregulation of this homeostasis is implicated in tumorigenesis and acquired resistance to treatments. The emerging importance of Mcl-1 protein in chemotherapeutic resistance makes it a high priority therapeutic target. Targeting PPIs associated with Mcl-1 presents many challenges for the design of inhibitors. This review focuses on the characterization of the Mcl-1 hot-spots which are related to four hydrophobic pockets P1-P4 and one major electrostatic interaction. Analysis of structural data highlights the high importance of the P2/P3 pockets for the binding of nonpeptide ligands. In order to guide medicinal chemists into making more selective and potent Mcl-1 inhibitors, the Mcl-1 protein is compared to other antiapoptotic proteins.

Early-life represents a critically sensitive window to endocrine disrupting chemicals, potentially leading to long-term repercussions on the phenotype later in life. The mechanisms underlying this phenomenon, referred to as the Developmental Origins of Health and Disease (DOHaD), are still poorly understood. To gain molecular understanding of these effects, we exposed mangrove rivulus (Kryptolebias marmoratus) for 28 days post hatching (dph) to 4 and 120 ng/L 17-α-ethinylestradiol, a model xenoestrogen. After 28 days, fish were raised for 140 days in clean water and we performed quantitative label-free proteomics on brain, liver and ovotestis of 168 dph adults. A total of 820, 888 and 420 proteins were robustly identified in the brain, liver and ovotestis, respectively. Effects of 17-α-ethinylestradiol were tissue and dose-dependent: a total of 31, 51 and 18 proteins were differentially abundant at 4 ng/L in the brain, liver and ovotestis, respectively, compared to 20, 25 and 39 proteins at 120 ng/L. Our results suggest that estrogen-responsive pathways, such as lipid metabolism, inflammation, and the innate immune system were affected months after the exposure. In addition, the potential perturbation of S-adenosylmethionine metabolism encourages future studies to investigate the role of DNA methylation in mediating the long-term effects of early-life exposures. Significance The Developmental Origins of Health and Disease (DOHaD) states that early life stages of humans and animals are sensitive to environmental stressors and can develop health issues later in life, even if the stress has ceased. Molecular mechanisms supporting DOHaD are still unclear. The mangrove rivulus is a new fish model species naturally reproducing by self-fertilization, making it possible to use isogenic lineages in which all individuals are highly homozygous. This species therefore permits to strongly reduce the confounding factor of genetic variability in order to investigate the effects of environmental stress on the phenotype. After characterizing the molecular phenotype of brain, liver and ovotestis, we obtained true proteomic reaction norms of these three organs in adults after early life stages have been exposed to the common endocrine disruptor 17-α-ethinylestradiol (EE2). Our study demonstrates long-term effects of early-life endocrine disruption at the proteomic level in diverse estrogen-responsive pathways 5 months after the exposure. The lowest tested and environmentally relevant concentration of 4 ng/L had the highest impact on the proteome in brain and liver, highlighting the potency of endocrine disruptors at low concentrations.

A woman’s nutritional status during pregnancy and breastfeeding is not only critical for her health, but also for that of future generations. Nutritional requirements during pregnancy differ considerably from those of non-pregnant women. Thus, a personalized approach to nutritional advice is recommended. Currently, some countries recommend routine supplementation for all pregnant women, while others recommend supplements only when necessary. Maternal physiological adaptations, as well as nutritional requirements during pregnancy and lactation, will be reviewed in the literature examining the impacts of dietary changes. All of these data have been studied deeply to facilitate a discussion on dietary supplement use and the recommended doses of nutrients during pregnancy and lactation. The aim of this review is to evaluate the knowledge in the scientific literature on the current recommendations for the intake of the most common micronutrients and omega-3 fatty acids during pregnancy and lactation in the United States, Canada, and Europe. Taking into account these considerations, we examine minerals, vitamins, and omega-3 fatty acid requirements. Finally, we conclude by discussing the potential benefits of each form of supplementation.

International audience; Protein-protein interactions are attractive targets because they control numerous cellular processes. In oncology, apoptosis regulating Bcl-2 family proteins are of particular interest. Apoptotic cell death is controlled via PPIs between the anti-apoptotic proteins hydrophobic groove and the pro-apoptotic proteins BH3 domain. In ovarian carcinoma, it has been previously demonstrated that Bcl-x L and Mcl-1 cooperate to protect tumor cells against apoptosis. Moreover, Mcl-1 is a key regulator of cancer cell survival and is a known resistance factor to Bcl-2/Bcl-x L pharmacological inhibitors making it an attractive therapeutic target. Here, using a structure-guided design from the oligopyridine lead Pyr-idoclax based on Noxa/Mcl-1 interaction we identified a new derivative, active at lower concentration as compared to Pyridoclax. This new derivative selectively binds to the Mcl-1 hydrophobic groove and releases Bak and Bim from Mcl-1 to induce cell death and sensitize cancer cells to Bcl-2/Bcl-x L targeting strategies.

International audience; AimsA better understanding of how plant growth, N nutrition and symbiotic nitrogen fixation (SNF) are influenced by soil inorganic N availability, for a wide range of legume species, is crucial to optimise legume productivity, N2 fixation, while limiting environmental risks such as N leaching. MethodsA comparative analysis was performed for ten legume crops, grown in a field experiment and supplied with four N fertiliser rates. Dry matter, N concentration and SNF were measured. In parallel, root elongation rates were studied in a greenhouse experiment. ResultsFor most species, N fertilisation had little effect on plant growth and N accumulation. SNF was reduced by soil inorganic N available at sowing but with large differences in the magnitude of the response among species. The response varied according to plant N requirements for growth and plant ability to retrieve inorganic N. Accordingly, root lateral expansion rate measured in RhizoTubes was highly correlated with plant ability to retrieve inorganic N measured in the field experiment. Conclusion Combining SNF response to soil inorganic N, shoot N and plant ability to retrieve inorganic N, allowed a robust evaluation of differential response to soil inorganic N among a wide range of legume species.

International audience; Cereal-legume intercropping is known to improve the sustainability of crop production. However, it remains uncommon on commercial farms in Europe due to a number of socio-technical lock-ins and the many practical issues raised when integrating intercrops in cropping systems (e.g. which species, cultivars, sowing densities). Crop modelling is an option to explore integration scenarios and support farmers' decisions. However, available crop models are not able to simulate bundles of ecosystem services provided by a large diversity of binary cereallegume intercropping scenarios. To address this challenge, we developed a hybrid modelling chain that combines process-based, statistical and knowledge-based models to benefit from the strengths of these three different modelling approaches. The chain (i) simulates potential biomass of the sole cereal and legume crops independently using the crop model STICS; (ii) uses statistical interaction models built in R to convert potential biomass in sole cropping into attainable biomass in intercropping by considering competition effects among species, using a field trial database; (iii) converts attainable biomass into actual biomass by considering pest damage using a knowledge-based multi-attribute DEXi model, and also assesses control of pests (i.e. weeds, insects and diseases); and (iv) uses another set of multi-attribute models to assess five additional ecosystem services (i.e. cereal and legume grain yields, cereal protein content, nitrogen supply to the following crop and impact on soil structure) from the actual biomass of the intercrop at harvest and/or cropping system features. The chain was calibrated for grain cereal-legume intercrops sown simultaneously in a random pattern under low-input French conditions. We used an expert-based approach to assess the performances of each model and evaluate the accuracy of the entire modelling chain. In 18 simulated scenarios, 79% of the predicted levels of ecosystem services were consistent with experts' opinion. Predictions were more accurate for intercropping scenarios that included species from the trial database used to build linear interaction models (relative RMSE of 27-31%) but remained satisfactory for other intercropped species (relative RMSE of 32-37%). This is the first modelling chain able to assess bundles of ecosystem services provided by multiple cereal-legume intercrops in function of their cropping system contexts. This chain is intended to be included in an educational tool that is used face to face with farmers or students to design cropping systems that include intercrops.

Cet article propose une analyse historique du processus de verrouillage du système agroalimentaire en défaveur des légumineuses à graines, à l’aune des théories évolutionnistes. Plusieurs mécanismes d’autorenforcement permettent de comprendre pourquoi ces espèces sont de moins en moins cultivées en France face à un système agro-industriel qui s’est spécialisé en faveur des céréales, favorisant à l’amont l’usage d’engrais azotés de synthèse et limitant à l’aval les investissements pour les légumineuses en alimentation humaine. Cet article s’interroge alors sur les perspectives de déverrouillage.

Protein-protein interactions (PPIs) control many important physiological processes within human cells. Apoptosis or programmed cell death is closely regulated by pro- and antiapoptotic signals. Dysregulation of this homeostasis is implicated in tumorigenesis and acquired resistance to treatments. The emerging importance of Mcl-1 protein in chemotherapeutic resistance makes it a high priority therapeutic target. Targeting PPIs associated with Mcl-1 presents many challenges for the design of inhibitors. This review focuses on the characterization of the Mcl-1 hot-spots which are related to four hydrophobic pockets P1-P4 and one major electrostatic interaction. Analysis of structural data highlights the high importance of the P2/P3 pockets for the binding of nonpeptide ligands. In order to guide medicinal chemists into making more selective and potent Mcl-1 inhibitors, the Mcl-1 protein is compared to other antiapoptotic proteins.

Protein–protein interactions (PPIs) are involved in many cellular processes; consequently, the discovery of small molecules as modulators of PPIs has become an important challenge in medicinal chemistry. Structural mimetics of α-helices, β-turns or β-strands could maintain or restore biological functions and should possess biological activity. At this time, the most challenging classes of PPIs are those mediated by β-sheet interactions, which are implicated in a number of diseases. Only a few β-strand mimics have been published to date. This study presents an evaluation of oligothienylpyridyl scaffolds in view of their ability for β-strand mimicry. In this study, theoretical ring twist angle predictions for these scaffolds have been validated by X-ray diffraction and molecular dynamics simulations with NMR constraints. Careful choice of substituent and heavy-atom positions in the foldamer units opens the way to produce reasonably coplanar compounds mimicking β-strand side-chain distribution.

International audience; Grain-legume plants fix atmospheric nitrogen in the soil and thus do not need nitrogen fertilizers. Therefore, grain- legumes can potentially decrease global warming, as nitrogen fertilization is responsible for half of all agricultural greenhouse gas emissions. Moreover, grain-legumes have many functional and nutritional properties both as feed and food. Despite the fact that the European Union has granted considerable subsidies to promote grain-legume cultivation, their production continues to fall and there has been no satisfactory explanation as to why. This study provides an answer by showing that a situation of technological lock-in has resulted from the co-evolution of crop systems, based on an agrochemical paradigm, public policies, and market dynamics that promote cereals. This process began with the historical choice by European and French public institutions to relegate grain-legumes to feed in direct competition with imported soybeans. Moreover, interrelated factors, such as breeding selection, public subsidies, and food systems, have favored increasing returns to adoption for cereals to the detriment of grain-legumes. Finally, the evolutionary economics approach used here identified several actions that must be implemented together, such as agricultural cost-accounting methods, nitrogen management, institutional innovations, and market out lets to promote grain-legumes and move towards more sustainable agriculture.

Myeloid cell leukemia-1 (Mcl-1) is an anti-apoptotic member of the Bcl-2 family proteins. Its amplification is one of the most frequent genetic aberrations found in human cancers. Pyridoclax, a promising BH3 mimetic inhibitor, interacts directly with Mcl-1 and induces massive apoptosis at a concentration of 15 µM in combination with anti-Bcl-xL strategies in chemo-resistant ovarian cancer cell lines. In this study, a combined experimental and theoretical approach was used to investigate the binding mode of Pyridoclax to Mcl-1. The representative poses generated from dynamics simulations compared with NMR data revealed: (i) Pyridoclax bound to P1 and P2 pockets of Mcl-1 BH3 binding groove through its styryl and methyl groups establishing mainly hydrophobic contacts, (ii) one of the ending pyridines interacts through electrostatic interaction with K234 side chain, a negatively charged residue present only in this position in Mcl-1. Communicated by Ramaswamy H. Sarma

In addition to genetic variation, epigenetic mechanisms such as DNA methylation might make important contributions to heritable phenotypic diversity in populations. However, it is often difficult to disentangle the contributions of genetic and epigenetic variation to phenotypic diversity. Here, we investigated global DNA methylation and mRNA expression of the methylation‐associated enzymes during embryonic development and in adult tissues of one natural isogenic lineage of mangrove rivulus fish, Kryptolebias marmoratus. Being the best‐known self‐fertilizing hermaphroditic vertebrate affords the opportunity to work with genetically identical individuals to examine, explicitly, the phenotypic effects of epigenetic variance. Using the LUminometric Methylation Assay (LUMA), we described variable global DNA methylation at CpG sites in adult tissues, which differed significantly between hermaphrodite ovotestes and male testes (79.6% and 87.2%, respectively). After fertilization, an immediate decrease in DNA methylation occurred to 15.8% in gastrula followed by re‐establishment to 70.0% by stage 26 (liver formation). Compared to zebrafish, at the same embryonic stages, this reprogramming event seems later, deeper, and longer. Furthermore, genes putatively encoding DNA methyltransferases (DNMTs), Ten‐Eleven Translocation (TET), and MeCP2 proteins showed specific regulation in adult gonad and brain, and also during early embryogenesis. Their conserved domains and expression profiles suggest that these proteins play important roles during reproduction and development. This study raises questions about mangrove rivulus’ peculiar reprogramming period in terms of epigenetic transmission and physiological adaptation of individuals to highly variable environments. In accordance with the general‐purpose genotype model, epigenetic mechanisms might allow for the expression of diverse phenotypes among genetically identical individuals. Such phenotypes might help to overcome environmental challenges, making the mangrove rivulus a valuable vertebrate model for ecological epigenetic studies. The mangrove rivulus, Kryptolebias marmoratus, is the best‐known self‐fertilizing hermaphroditic vertebrate that allows to work with genetically identical individuals to examine, explicitly, the phenotypic effects of epigenetic variance. The reprogramming event is later, more dramatic and longer than in other described vertebrates. High evolutionary conservation and expression patterns of DNMT, TET, and MeCP2 proteins in K. marmoratus suggest biological roles for each member in gametogenesis and development.

International audience; Mcl-1, which is an anti-apoptotic member of the Bcl-2 protein family, is overexpressed in various cancers and promotes the aberrant survival of tumor cells. To inhibit Mcl-1, and initiate apoptosis, an interaction between BH3-only proteins and Mcl-1 anti-apoptotic protein is necessary. These protein-protein interactions exhibit some selectivity: Mcl-1 binds specifically to Noxa, whereas Bim and Puma bind strongly to all anti-apoptotic proteins. Even if the three-dimensional (3D) structures of several Mcl-1/BH3-only complexes have been solved, the BH3-only binding specificity to Mcl-1 is still not completely understood. In this study, molecular dynamics simulations were used to elucidate the molecular basis of the interactions with Mcl-1. Our results corroborate the importance of four conserved hydrophobic residues and a conserved aspartic acid on BH3-only as a common binding pattern. Furthermore, our results highlight the contribution of the fifth hydrophobic residue in the C-terminal part and a negatively charged patch in the N-terminal of BH3-only peptides as important for their fixation to Mcl-1. We hypothesize that this negatively charged patch will be an Mcl-1 specific binding pattern

International audience; Alzheimer's Disease (AD) is a neurodegenerative brain disorder in which many biological dysfunctions are involved. Among them, two main types of lesions were discovered and widely studied: the amyloid plaques and the neurofibrillary tangles (NFTs). These two lesions are caused by the dysfunction and the accumulation of two proteins which are, respectively, the beta-amyloid peptide and the tau protein. The process that leads these two proteins to aggregate is complex and is the subject of current studies. After a brief description of the aggregation mechanisms, we will provide an overview of new therapeutic agents targeting the different dysfunctions and toxic species found during aggregation

Triclosan (TCS) is a broad spectrum antibacterial agent widely used in personal care products and present in most aquatic ecosystems. This study investigated the occurrence of triclosan acclimation and the biological mechanisms underlying the stress response triggered in early-life stage of zebrafish. Zebrafish eggs were first exposed to four different sublethal concentrations of TCS (2, 20, 50 and 100 μg/L) for 7 days following fertilization and subsequently exposed to a lethal concentration of TCS (1000 μg/L). During the time-to-death exposure (TTD), mortality was continuously recorded to evaluate if increased resistance occurred. Overall, larvae exposed to 50 μg/L of TCS demonstrated higher sensitivity, with delayed hatching and increased mortality during the sub-lethal exposure and significant lower mean time-to-death (TTD) value compared to the other groups. Interestingly, fish exposed to the highest concentration of TCS (100 μg/L) presented a similar mean TTD value as controls and a significantly better survival in comparison with embryos exposed to 50 μg/L, suggesting that acclimation process has been triggered at this concentration. Proteomic and enzymatic analyses were conducted on 7 days post fertilization (dpf) larvae exposed to 50 μg/L and 100 μg/L of TCS giving insights into the functional changes triggered at those specific concentrations. TCS seemed to affect proteins involved in cytoskeleton, stress response, eyes and neuronal development. This was endorsed by the enzymatic results, which suggest impairment in glutathione metabolism and acute neurotoxicity. A significant 2.5-fold and 3-fold increase of AChE activity was observed following TCS exposure. Moreover, GPx activity was significantly increased whereas a significant inhibition of GR activity was observed, suggesting that de novo synthesis of reduced GSH might occur in order to maintain the ratio between reduced and oxidized GSH. Proteomic results revealed possible candidate protein involved in the acclimation process of larvae exposed to 100 μg/L of TCS. Our integrative analysis revealed complex non-monotonic concentration-related effects on zebrafish early-life stages with increased resistance between 50 and 100 μg/L exposures. This research highlighted oxidative stress and neurotoxicity as major toxicity mechanisms of TCS during development.

International audience; Overexpression of efflux pumps is an important mechanism of bacterial resistance that results in the extrusion of antimicrobial agents outside the bacterial cell. Inhibition of such pumps appears to be a promising strategy that could restore the potency of existing antibiotics. The NorA efflux pump of Staphylococcus aureus confers resistance to a wide range of unrelated substrates, such as hydrophilic fluoroquinolones, leading to a multidrug-resistance phenotype. In this work, approximately 150 heterocyclic boronic species were evaluated for their activity against susceptible and resistant strains of S. aureus. Twenty-four hit compounds, although inactive when tested alone, were found to potentiate ciprofloxacin activity by a 4-fold increase at concentrations ranging from 0.5 to 8 μg/mL against S. aureus 1199B, which overexpresses NorA. Boron-free analogues showed no biological activity, thus revealing that the boron atom is crucial for biological activity. This work describes the first reported efflux pump inhibitory activity of boronic acid derivatives.

International audience; As a legume, pea plant has the ability to symbiotically fix N-2. However, symbiotic N-2 fixation is very sensitive to environmental stresses that affect plant growth, and there is little knowledge on the impact of root pruning on N-2 fixation and plant growth. In this study, we removed half of the nodulated roots of pea wild-type Frisson and hypernodulating mutants P64, P118, and P121. Dinitrogen fixation was measured using N-15 labeling and carbon assimilation and partitioning between plant organs using C-13 labeling. Root pruning decreased N-2 fixation by -46 to -79 % in wild-type and mutants. Pea mutant P118 had a lower decrease of specific activity of N-2 fixation (-17 %) than both wild-type and other mutants (-36 to -62 %). For all genotypes, root pruning increased root and nodule sinks strengths for carbon. For P118 and for P121, this was associated to higher nodule growth than for control plants, as measured 8 days after root pruning. This is the first analysis of N-2-fixing plant response to root pruning. Importantly, we showed that some hypernodulating mutant pea lines (P118 and to a lesser extent P121) withstood this stress better than wild-type did.

Protein–protein interactions (PPIs) play a central role in all biological processes and have been the focus of intense investigations from structural molecular biology to cell biology for the majority of the last two decades and, more recently, are emerging as important targets for pharmaceuticals. A common motif found at the interface of PPIs is the α-helix, and apart from the peptidic structures, numerous nonpeptidic small molecules have been developed to mimic α-helices. The first-generation terphenyl scaffold is able to successfully mimic key helix residues and disrupt relevant interactions, including Bcl-xL-Bak interactions that are implicated in apoptosis mechanism. These scaffolds were designed and evaluated in silico. Analysis revealed that substituents on aromatic scaffolds can efficiently mimic side-chain surfaces. Unfortunately, the literature describes a long and difficult procedure to access these aromatic-based scaffolds. The search for new simpler methodology is the aim of the research of our medicinal chemistry team. On the basis of structural requirements, we developed a program concerning the synthesis of new oligo(het)aryl scaffolds produced by iterative couplings of boronic species (garlanding) in which substituents on rings project functionality in spatial orientations that mimic residues of an α-helix.

Proteins modulate the majority of all biological functions and are composed of highly organized secondary structural elements such as helices, turns, and sheets. Many of these functions are affected by a small number of key structural element, protein–protein interactions. Their mimicry by peptide and non-peptide scaffolds has become a major focus of contemporary research. This paper examines oligomeric system as new foldamers, which either reproduce the local topography of the helix, or project appropriately functionality in a similar manner to residues of an alpha-helix.

This paper describes for the first time an efficient approach of Suzuki Miyaura cross-coupling reaction leading to new mixed linear unsymmetric phenylpyridyl chains (garlands). We have studied the synthesis of new phenylpyridyl boronic species and their reactivity with a dihalogenated bipyridine to obtain four and six unit phenylpyridines.

This paper describes an efficient route leading to new thienylpyridyl garlands as non-peptidic alpha helix mimetics and potential protein–protein interactions disruptors. Firstly, we have studied the reactivity of boronic acids and halogenated pyridines and/or thiophenes towards the Suzuki–Miyaura cross-coupling reaction in order to obtain bis-thienylpyridines. Secondly, we have functionalized these compounds by a reaction of bromination and the resultant bis-bromothienylpyridines have been found to undergo iterative Pd-catalyzed coupling based on a pseudo-Garlanding approach with a range of pyridyl boronic acids to produce a new library of thienylpyridyl oligomers.

Ovarian cancer is the leading cause of death from gynecological malignancies worldwide. Although the patients are initially quite sensitive to the taxane and platinum-based first-line chemotherapy, most of them relapse and develop chemoresistance. Defects in apoptosis regulation in ovarian cancer allow the cancer cells to evade cell death and contribute to chemoresistance. Mcl-1 is an anti-apoptotic member of the Bcl-2 proteins family and its amplifıcation is one of the most frequent genetic aberrations found in human cancers. Its expression is at the origin of the acquired resistance to chemotherapy and to Bcl-2 and Bcl-xL inhibitors. In ovarian cancers, we previously demonstrated that Bcl-xL and Mcl-1 cooperate to prevent cancer cells from undergoing apoptotic cell death. Their concomitant inhibitions lead to massive apoptosis even in absence of chemotherapy. Moreover, in some cases, Mcl-1 inhibition is itself able to lead to apoptosis. If clinically relevant pharmacologic inhibition of Bcl-xL is available using ABT-263, selective direct inhibition of Mcl-1 remains problematic. In this context, our teams have designed and synthesized small compounds based on a pyridyl scaffold, named oligopyridines, which potentially target the Mcl-1 hydrophobic binding pocket. We demonstrated that the lead of the first generation of oligopyridines, named Pyridoclax, interacts directly with Mcl-1, releases its pro-apoptotic partners Bim and Bak and induces massive apoptosis at 25 µM concentration in combination with anti-Bcl-xL strategies in chemoresistant ovarian cancer cell lines (Gloaguen et al., J Med Chem 2015). In the present study, we investigated the antitumor activity of Pyridoclax hydrochloride in three subcutaneous xenograft models derived from the injection of chemoresistant ovarian cancer cell lines. Different routes of Pyridoclax hydrochloride administration were tested and its antitumor effect was analyzed at different doses as single agent or in combination with ABT-263. This study highlighted an effective antitumor activity of 20mg/kg of Pyridoclax administered intravenously as single agent in two of three xenograft models without side effects. In order to improve its biological activity, we evaluated the cytotoxic effects of a second generation of oligopyridines derived from the Pyridoclax. This allowed us to identify the MR31367, one of the most potent oligopyridines that shows a stronger pro-apoptotic activity in association with to Bcl-xL-targeting strategies in ovarian cancer cell lines. Further characterization showed that this derivative binds Mcl-1 and release Bim and Bak from it, leading to Bak-mediated apoptosis. Overall, these results open up interesting perspectives for the clinical use of Mcl-1 inhibitors as single agent or in combination with anticancer drugs to improve the clinical management of ovarian cancers. Citation Format: Siham Hedir, Louis-Bastien Weiswald, Marcella De Giorgi, Jade Fogha, Martina De Pascale, Emilie Brotin, Bogdan Marekha, Peggy Suzanne, Fabien Gautier, Philippe Juin, Laetitia Ligat, Frédéric Lopez, Rémi Legay, Ronan Bureau, Sylvain Rault, Jana Sopkova-de Oliveira Santos, Anne-Sophie Voisin-Chiret, Laurent Poulain. Pyridoclax and its derivatives from oligopyridine family directly inhibit Mcl-1 and exert potent antitumor effects on ovarian cancer in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3996.

The identification of novel therapeutic strategies is an important urgent requirement for the clinical management of ovarian cancers, which remain the leading cause of death from gynecologic cancer. Our previous studies have shown that the anti-apoptotic proteins Bcl-xL and Mcl-1 cooperate to protect resistant ovarian cancer cells from apoptosis as their concomitant inhibition results in massive apoptotic cell death. Moreover, their BH3-only pro-apoptotic partners Bim and Puma are crucial actors in the induced cell death. This suggests that ovarian cancer cell apoptosis can be triggered by inhibiting Bcl-xL and Mcl-1 and/or by promoting their BH3-only partners, especially Bim and Puma. The expression of these Bcl-2 family proteins has been reported to be modulated by HDAC (histone deacetylases) inhibitors in various cancer models. The objective of the present study was to evaluate the efficacy of the FDA-approved pan-HDAC inhibitor belinostat (i) to reduce the [Bcl-xL and Mcl-1] / BH3-only proteins ratio and (ii) to trigger apoptosis in chemoresistant ovarian cancer cell lines, alone or in combination with other pharmacological molecules that modulate these Bcl-2 family proteins. HDAC inhibition by belinostat led to a G2/M blockade and consequently reduced cell proliferation. Moreover, belinostat used at high concentrations (from 1000 to 2000 nM) efficiently triggered apoptosis in chemoresistant ovarian cancer cells. This cytotoxic effect was associated with both an up-regulation of Bim and Puma protein expression and an inhibition of Bcl-xL protein expression. Used at sub-toxic concentrations, belinostat also up-regulated Bim and Puma proteins but failed to down-regulate Bcl-xL and Mcl-1 protein expression. We therefore investigated whether Bcl-xL or Mcl-1 inhibition could sensitize ovarian cancer cells to sub-toxic concentrations of belinostat. Interestingly, the combination of belinostat with the BH3-mimetic ABT-737, that inhibits Bcl-xL, induced massive apoptosis. Moreover, Mcl-1 direct or indirect inhibition was also very efficient to sensitize ovarian cancer cells to belinostat. These results suggest that belinostat, alone or in association with pharmacological molecules that inhibit Mcl-1 or Bcl-xL, represents a promising approach for the treatment of chemoresistant ovarian cancers. Citation Format: Charlène Duboc, Edwige Abeilard, Anne Sophie Voisin-Chiret, Pascal Gauduchon, Laurent Poulain, Marie Villedieu. The sensitivity of ovarian cancer cells to the HDAC inhibitor belinostat is improved by inhibiting Bcl-xL or Mcl-1 anti-apoptotic proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1921.

This paper describes an efficient and regioselective synthetic route leading to new dihalobi- and terpyridines. We developed a strategy based on regioselective sequence of Suzuki–Miyaura cross-coupling reactions between bromopyridyl boronic acids and dihalopyridines and dihalobipyridines. The study of the influence of the nature and the position of the halogen atoms leads to prepare bromoiododerivatives to obtain good selectivities.

This paper sets forth a global synthetic strategy based on the Garlanding concept to design and to produce halo-oligopyridines. This strategy allows to prepare an infinite number of these new compounds and hence to create a library of halo-oligopyridines. This article is focused on the basic principle of the Garlanding concept and on the demonstration of its feasibility.

A synthetic method for the preparation of 6-aryl-1,4-dimethyl-9H-carbazoles involving a palladium catalyzed coupling reaction of 1,4-dimethyl-9H-carbazole-6-boronic acids and (hetero)aryl halides is described.

This paper sets forth the synthesis of novel oxybispyridylboronic acids, which are prepared from the corresponding halo-oxybispyridines via halogen–metal exchange using n-butyllithium and treatment with triisopropylborate. A range of efficient cross-coupling reactions of these novel boronic acids with selected aryl halides is described. This strategy produces novel pyridylethers of interest in cholinergic medicinal chemistry.

The implementation of the Petasis boronic Mannich reaction in pyridine series allowed us to obtain original compounds whose structure was investigated and determined a stable complex (1:1) of dioxaborolanone and amine.

The effect of nitrogen source (N(2) or nitrate) on carbon assimilation by photosynthesis and on carbon partitioning between shoots and roots was investigated in pea (Pisum sativum L. 'Baccara') plants at different growth stages using (13)C labelling. Plants were grown in the greenhouse on different occasions in 1999 and 2000. Atmospheric [CO(2)] and growth conditions were varied to alter the rate of photosynthesis. Carbon allocation to nodulated roots was unaffected by N source. At the beginning of the vegetative period, nodulated roots had priority for assimilates over shoots; this priority decreased during later stages and became identical to that of the shoot during seed filling. Carbon allocation to nodulated roots was always limited by competition with shoots, and could be predicted for each phenological stage: during vegetative and flowering stages a single, negative exponential relationship was established between sink intensity (percentage of C allocated to the nodulated root per unit biomass) and net photosynthesis. At seed filling, the amount of carbon allocated to the nodulated root was directly related to net photosynthesis. Respiration of nodulated roots accounted for more than 60 % of carbon allocated to them during growth. Only at flowering was respiration affected by N supply: it was significantly higher for strictly N(2)-fixing plants (83 %) than for plants fed with nitrate (71 %). At the vegetative stage, the increase in carbon in nodulated root biomass was probably limited by respiration losses.

International audience; Apoptosis control defects such as the deregulation of Bcl-2 family member expression are frequently involved in chemoresistance. In ovarian carcinoma, we previously demonstrated that Bcl-xL and Mcl-1 cooperate to protect cancer cells against apoptosis and their concomitant inhibition leads to massive apoptosis even in the absence of chemotherapy. Whereas Bcl-xL inhibitors are now available, Mcl-1 inhibition, required to sensitize cells to Bcl-xL-targeting strategies, remains problematic. In this context, we designed and synthesized oligopyridines potentially targeting the Mcl-1 hydrophobic pocket, evaluated their capacity to inhibit Mcl-1 in live cells, and implemented a functional screening assay to evaluate their ability to sensitize ovarian carcinoma cells to Bcl-xL-targeting strategies. We established structure–activity relationships and focused our attention on MR29072, named Pyridoclax. Surface plasmon resonance assay demonstrated that pyridoclax directly binds to Mcl-1. Without cytotoxic activity when administered as a single agent, pyridoclax induced apoptosis in combination with Bcl-xL-targeting siRNA or with ABT-737 in ovarian, lung, and mesothelioma cancer cells

The influence of soil nitrate availability, crop growth rate and phenology on the activity of symbiotic nitrogen fixation (SNF) during the growth cycle of pea (Pisum sativum cv. Baccara) was investigated in the field under adequate water availability, applying various levels of fertiliser N at the time of sowing. Nitrate availability in the ploughed layer of the soil was shown to inhibit both SNF initiation and activity. Contribution of SNF to total nitrogen uptake (%Ndfa) over the growth cycle could be predicted as a linear function of mineral N content of the ploughed layer at sowing. Nitrate inhibition of SNF was absolute when mineral N at sowing was over 380 kg N ha−1. Symbiotic nitrogen fixation was not initiated unless nitrate availability in the soil dropped below 56 kg N ha−1. However, SNF could no longer be initiated after the beginning of seed filling (BSF). Other linear relationships were established between instantaneous %Ndfa and instantaneous nitrate availability in the ploughed layer of the soil until BSF. Instantaneous %Ndfa decreased linearly with soil nitrate availability and was nil above 48 and 34 kg N ha−1 for the vegetative and reproductive stages, respectively, levels after which no SNF occurred. Moreover, SNF rate was shown to be closely related to the crop growth rate until BSF. The ratio of SNF rate over crop growth rate decreased linearly with thermal time. Maximum SNF rate was about 40 mg N m−2 degree-day−1, equivalent to 7 kg N ha−1, regardless of the N treatment. From BSF to the end of the growth cycle, the high N requirements of the crop were supported by both SNF and nitrate root absorption but, of the two sources, nitrate root absorption seemed to be less affected by the presence of reproductive organs. However, since soil nitrate availability was low at the end of the growth cycle, SNF was the main source of nitrogen acquisition. The onset of SNF decrease at the end of the growth cycle seemed to be first due to nodule age and then associated to the slowing of the crop growth rate.

The effect of mineral N availability on nitrogen nutrition and biomass partitioning between shoot and roots of pea (Pisum sativum L., cv Baccara) was investigated under adequately watered conditions in the field, using five levels of fertiliser N application at sowing (0, 50, 100, 200 and 400 kg N ha−1). Although the presence of mineral N in the soil stimulated vegetative growth, resulting in a higher biomass accumulation in shoots in the fertilised treatments, neither seed yield nor seed nitrogen concentration was affected by soil mineral N availability. Symbiotic nitrogen fixation was inhibited by mineral N in the soil but it was replaced by root mineral N absorption, which resulted in optimum nitrogen nutrition for all treatments. However, the excessive nitrogen and biomass accumulation in the shoot of the 400 kg N ha−1 treatment caused crop lodging and slightly depressed seed yield and seed nitrogen content. Thus, the presumed higher carbon costs of symbiotic nitrogen fixation, as compared to root mineral N absorption, affected neither seed yield nor the nitrogen nutrition level. However, biomass partitioning within the nodulated roots was changed. The more symbiotic nitrogen fixation was inhibited, the more root growth was enhanced. Root biomass was greater when soil mineral N availability was increased: root growth was greater and began earlier for plants that received mineral N at sowing. Rooting density was also promoted by increased mineral N availability, leading to more numerous but finer roots for the fertilised treatments. However, the maximum rooting depth and the distribution of roots with depth were unchanged. This suggested an additional direct promoting effect of mineral N on root proliferation.

In order to highlight our understanding on ecosystems functioning and resource sharing/competition, either in artificial environment or agrosystems, according to changes in the climatic conditions, it is necessary to measure accurately element fluxes within plants. Stable isotopes allow tracking safely and accurately on a short time frame the behavior of elements in plants. After a short review devoted to isotopic studies of elemental flux within plants, we explain how a direct multiple labelling study might be conducted in a plant, so as to measure over short time nitrogen and sulfur acquisition, and assimilates arising from a labelled source.

International audience; Plant and soil types are usually considered as the two main drivers of the rhizosphere microbial communities. The aim of this work was to study the effect of both N availability and plant genotype on the plant associated rhizosphere microbial communities, in relation to the nutritional strategies of the plant-microbe interactions, for six contrasted Medicago truncatula genotypes. The plants were provided with two different nutrient solutions varying in their nitrate concentrations (0 mM and 10 mM). First, the influence of both nitrogen availability and Medicago truncatula genotype on the genetic structure of the soil bacterial and fungal communities was determined by DNA fingerprint using Automated Ribosomal Intergenic Spacer Analysis (ARISA). Secondly, the different nutritional strategies of the plant-microbe interactions were evaluated using an ecophysiological framework. We observed that nitrogen availability affected rhizosphere bacterial communities only in presence of the plant. Furthermore, we showed that the influence of nitrogen availability on rhizosphere bacterial communities was dependent on the different genotypes of Medicago truncatula. Finally, the nutritional strategies of the plant varied greatly in response to a modification of nitrogen availability. A new conceptual framework was thus developed to study plant-microbe interactions. This framework led to the identification of three contrasted structural and functional adaptive responses of plant-microbe interactions to nitrogen availability.

This paper describes an efficient route leading to new thienylpyridyl garlands as non-peptidic alpha helix mimetics and potential protein–protein interactions disruptors. Firstly, we have studied the reactivity of boronic acids and halogenated pyridines and/or thiophenes towards the Suzuki–Miyaura cross-coupling reaction in order to obtain bis-thienylpyridines. Secondly, we have functionalized these compounds by a reaction of bromination and the resultant bis-bromothienylpyridines have been found to undergo iterative Pd-catalyzed coupling based on a pseudo-Garlanding approach with a range of pyridyl boronic acids to produce a new library of thienylpyridyl oligomers.

One-pot reaction for the synthesis of novel phenylpyridyl derivatives and mixed quater phenylpyridyl compounds is described by using the Garlanding approach. The reactions proceed with moderate to good yields in mild conditions and good reaction times. This work represents a second application of the simplicity and versatility of Garlanding concept for the construction of new phenylpyridyl scaffolds, which can be considered as non-peptidic foldamer α-helix mimetics.

This paper describes an efficient and regioselective synthetic route leading to new dihalobi- and terpyridines. We developed a strategy based on regioselective sequence of Suzuki–Miyaura cross-coupling reactions between bromopyridyl boronic acids and dihalopyridines and dihalobipyridines. The study of the influence of the nature and the position of the halogen atoms leads to prepare bromoiododerivatives to obtain good selectivities.