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International audience; Designing appropriate management plans requires knowledge of both the dispersal ability and what has shaped the current distribution of the species under consideration. Here, we investigated the evolutionary history of the endangered gray reef shark (Carcharhinus amblyrhynchos) across its range by sequencing thousands of RADseq loci in 173 individuals in the Indo-Pacific (IP). We first bring evidence of the occurrence of a range expansion (RE) originating close to the Indo-Australian Archipelago (IAA) where two stepping-stone waves (east and westward) colonized almost the entire IP. Coalescent modeling additionally highlighted a homogenous connectivity (Nm ~ 10 per generation) throughout the range, and isolation by distance model suggested the absence of barriers to dispersal despite the affinity of C. amblyrhynchos to coral reefs. This coincides with long-distance swims previously recorded, suggesting that the strong genetic structure at the IP scale (FST ~ 0.56 between its ends) is the consequence of its broad current distribution and organization in a large number of demes. Our results strongly suggest that management plans for the gray reef shark should be designed on a range-wide rather than a local scale due to its continuous genetic structure. We further contrasted these results with those obtained previously for the sympatric but strictly lagoon-associated Carcharhinus melanopterus, known for its restricted dispersal ability. Carcharhinus melanopterus exhibits a similar RE dynamic but is characterized by a stronger genetic structure and a nonhomogeneous connectivity largely dependent on local coral reefs availability. This sheds new light on shark evolution, emphasizing the roles of IAA as source of biodiversity and of life-history traits in shaping the extent of genetic structure and diversity.

The tourism activities linked to artificial provisioning of blacktip reef sharks (Carcharhinus melanopterus) and pink whiprays (Pateobatis fai) on a specific site in French Polynesia were suddenly and completely stopped due to a COVID-19 lockdown that lasted 6 weeks from March 20 until April 30, 2020. Using both drone footage and underwater counting, we were able to track the abundance of those two species before, during, and after reopening and thus investigate the impact of provisioning on wild shark populations. The absence of any stimulus during this long period resulted in almost total desertion of the site by the elasmobranchs. However, 1 day prior to reopening, some individuals of both species positively reacted to the single acoustic stimulus of an engine boat, showing the resilience of conditioning, and some elasmobranchs reacted to acoustic and olfactive stimuli linked to the provisioning practice from the first day after reopening. During the first 2 weeks after reopening, the abundance of both species remained at reduced levels comparable to those observed between 2008 and 2010 for sharks;i.e., around 9 animals in the presence of local tourists. Pre-lockdown abundance levels, reaching approximatively 15 individuals for sharks and 10 for rays, were considered restored 1 and 2 months after reopening for blacktip reef sharks and pink whiprays, respectively. These findings improve our capacity to better understand the potential effects of artificial provisioning tourism on the abundance of elasmobranchs by showing that conditioning is resilient for several weeks, suggesting that intermittent interruption of elasmobranchs feeding would not really help to decrease its impact on animal welfare.

Knowledge about spatial and temporal variability in the distribution and abundance of predators is necessary to adapt measures to mitigate human–wildlife interactions. Acoustic telemetry and network analyses were used to investigate the spatial ecology of bull sharks, the species responsible for most shark bites in Reunion Island, one of the world's shark bite hotspots. The west coast of the island was not used uniformly by every individual, with size predicting the movements of sharks along the coast. Node-based metrics – closeness, node strength, and cumulated continuous residency times – derived from up to 181 monthly movement networks from 20 individuals, revealed that smaller sharks (

An animal's energy landscape considers the power requirements associated with residing in or moving through habitats. Within marine environments, these landscapes can be dynamic as water currents will influence animal power requirements and can change rapidly over diel and tidal cycles. In channels and along slopes with strong currents, updraft zones may reduce energy expenditure of negatively buoyant fishes that are also obligate swimmers. Despite marine predators often residing within high-current area, no study has investigated the potential role of the energetic landscape in driving such habitat selectivity. Over 500 grey reef sharks Carcharhinus amblyrhynchos reside in the southern channel of Fakarava Atoll, French Polynesia. We used diver observations, acoustic telemetry and biologging to show that sharks use regions of predicted updrafts and switch their core area of space use based on tidal state (incoming versus outgoing). During incoming tides, sharks form tight groups and display shuttling behaviour (moving to the front of the group and letting the current move them to the back) to maintain themselves in these potential updraft zones. During outgoing tides, group dispersion increases, swimming depths decrease and shuttling behaviours cease. These changes are likely due to shifts in the nature and location of the updraft zones, as well as turbulence during outgoing tides. Using a biomechanical model, we estimate that routine metabolic rates for sharks may be reduced by 10%-15% when in updraft zones. Grey reef sharks save energy using predicted updraft zones in channels and 'surfing the slope'. Analogous to birds using wind-driven updraft zones, negatively buoyant marine animals may use current-induced updraft zones to reduce energy expenditure. Updrafts should be incorporated into dynamic energy landscapes and may partially explain the distribution, behaviour and potentially abundance of marine predators.

Distinguishing the factors that influence activity within a species advances understanding of their behavior and ecology. Continuous observation in the marine environment is not feasible but biotelemetry devices provide an opportunity for detailed analysis of movements and activity patterns. This study investigated the detail that calibration of accelerometers measuring root mean square (RMS) acceleration with video footage can add to understanding the activity patterns of male and female Port Jackson sharks (Heterodontus portusjacksoni) in a captive environment. Linear regression was used to relate RMS acceleration output to time‐matched behavior captured on video to quantify diel activity patterns. To validate captive data, diel patterns from captive sharks were compared with diel movement data from free‐ranging sharks using passive acoustic tracking. The RMS acceleration data showed captive sharks exhibited nocturnal diel patterns peaking during the late evening before midnight and decreasing before sunrise. Correlation analysis revealed that captive animals displayed similar activity patterns to free‐ranging sharks. The timing of wild shark departures for migration in the late breeding season corresponded with elevated diel activity at night within the captive individuals, suggesting a form of migratory restlessness in captivity. By directly relating RMS acceleration output to activity level, we show that sex, time of day, and sex‐specific seasonal behavior all influenced activity levels. This study contributes to a growing body of evidence that RMS acceleration data are a promising method to determine activity patterns of cryptic marine animals and can provide more detailed information when validated in captivity.

Defects in the respiratory chain, interfering with energy production in the cell, are major underlying causes of mitochondrial diseases. In spite of this, the surprising variety of clinical symptoms, disparity between ages of onset, as well as the involvement of mitochondrial impairment in ageing and age-related diseases continue to challenge our understanding of the pathogenic processes. This complexity can be in part attributed to the unique metabolic needs of organs or of various cell types. In this view, it remains essential to investigate mitochondrial dysfunction at the cellular level. For this purpose, we developed a novel enzyme histochemical method that enables precise quantification in fresh-frozen tissues using competing redox reactions which ultimately lead to the reduction of tetrazolium salts and formazan deposition in cytochrome c oxidase-deficient mitochondria. We demonstrate that the loss of oxidative activity is detected at very low levels - this achievement is unequalled by previous techniques and opens up new opportunities for the study of early disease processes or comparative investigations. Moreover, human biopsy samples of mitochondrial disease patients of diverse genotypic origins were used and the successful detection of COX-deficient cells suggests a broad application for this new method. Lastly, the assay can be adapted to a wide range of tissues in the mouse and extends to other animal models, which we show here with the fruit fly, Drosophila melanogaster. Overall, the new assay provides the means to quantify and map, on a cell-by-cell basis, the full extent of COX deficiency in tissues, thereby expending new possibilities for future investigation. © 2018 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Mitochondrial dysfunction in adipose tissue occurs in obesity, type 2 diabetes, and some forms of lipodystrophy, but whether this dysfunction contributes to or is the result of these disorders is unknown. To investigate the physiological consequences of severe mitochondrial impairment in adipose tissue, we generated mice deficient in mitochondrial transcription factor A (TFAM) in adipocytes by using mice carrying adiponectin-Cre and TFAM floxed alleles. These adiponectin TFAM-knockout (adipo-TFAM-KO) mice had a 75–81% reduction in TFAM in the subcutaneous and intra-abdominal white adipose tissue (WAT) and interscapular brown adipose tissue (BAT), causing decreased expression and enzymatic activity of proteins in complexes I, III, and IV of the electron transport chain (ETC). This mitochondrial dysfunction led to adipocyte death and inflammation in WAT and a whitening of BAT. As a result, adipo-TFAM-KO mice were resistant to weight gain, but exhibited insulin resistance on both normal chow and high-fat diets. These lipodystrophic mice also developed hypertension, cardiac hypertrophy, and cardiac dysfunction. Thus, isolated mitochondrial dysfunction in adipose tissue can lead a syndrome of lipodystrophy with metabolic syndrome and cardiovascular complications.—Vernochet, C., Damilano, F., Mourier, A., Bezy, O., Mori, M. A., Smyth, G., Rosenzweig, A., Larsson, N.-G., Kahn, C. R. Adipose tissue mitochondrial dysfunction triggers a lipodystrophic syndrome with insulin resistance, hepatosteatosis, and cardiovascular complications.

During a survey of the population of blacktip reef shark Carcharhinus melanopterus in Moorea (French Polynesia) between 2007 and 2011, population structural characteristics were estimated from 268 individuals. Total length (L T ) ranged from 48 to 139 cm and 48 to 157 cm for males and females, respectively, demonstrating that the average L T of females was larger than that of males. The C. melanopterus population at Moorea showed an apparent spatial sexual segregation with females preferentially frequenting lagoons and males the fore-reefs. Mean growth rate was c. 6 cm year ―1 . Males reached sexual maturity at 111cm L T . This study reports on the population characteristics of this widespread carcharhinid shark species and makes comparisons with other locations, confirming high geographic variability in the population structure of the species.

BACKGROUND AND PURPOSE Muscarinic and adrenergic G protein-coupled receptors (GPCRs) are the targets of rare peptide toxins isolated from snake or cone snail venoms. We used a screen to identify novel toxins from Dendroaspis angusticeps targeting aminergic GPCRs. These toxins may offer new candidates for the development of new tools and drugs. EXPERIMENTAL APPROACH In binding experiments with 3H-rauwolscine, we studied the interactions of green mamba venom fractions with α2-adrenoceptors from rat brain synaptosomes. We isolated, sequenced and chemically synthesized a novel peptide, ρ-Da1b. This peptide was pharmacologically characterized using binding experiments and functional tests on human α2-adrenoceptors expressed in mammalian cells. KEY RESULTS ρ-Da1b, a 66-amino acid peptide stabilized by four disulphide bridges, belongs to the three-finger-fold peptide family. Its synthetic homologue inhibited 80% of 3H-rauwolscine binding to the three α2-adrenoceptor subtypes, with an affinity between 14 and 73 nM and Hill slopes close to unity. Functional experiments on α2A-adrenoceptor demonstrated that ρ-Da1b is an antagonist, shifting adrenaline activation curves to the right. Schild regression revealed slopes of 0.97 and 0.67 and pA2 values of 5.93 and 5.32 for yohimbine and ρ-Da1b, respectively. CONCLUSIONS AND IMPLICATIONS ρ-Da1b is the first toxin identified to specifically interact with α2-adrenoceptors, extending the list of class A GPCRs sensitive to toxins. Additionally, its affinity and atypical mode of interaction open up the possibility of its use as a new pharmacological tool, in the study of the physiological roles of α2-adrenoceptor subtypes.

Background information. The idea that GPCRs (G-protein-coupled receptors) may exist as homo- or hetero-oligomers, although still controversial, is now widely accepted. Nevertheless, the functional roles of oligomerization are still unclear and gaining greater insight into the mechanisms underlying the dynamics of GPCR assembly and, in particular, assessing the effect of ligands on this process seems important. We chose to focus our present study on the effect of MT7 (muscarinic toxin 7), a highly selective allosteric peptide ligand, on the oligomerization state of the hM1 (human M1 muscarinic acetylcholine receptor subtype). Results. We analysed the hM1 oligomerization state in membrane preparations or in live cells and observed the effect of MT7 via four complementary techniques: native-PAGE electrophoresis analysed by both Western blotting and autoradiography on solubilized membrane preparations of CHO-M1 cells (Chinese-hamster ovary cells expressing muscarinic M1 receptors); FRET (fluorescence resonance energy transfer) experiments on cells expressing differently tagged M1 receptors using either an acceptor photobleaching approach or a novel fluorescence emission anisotropy technique; and, finally, by BRET (bioluminescence resonance energy transfer) assays. Our results reveal that MT7 seems to protect the M1 receptor from the dissociating effect of the detergent and induces an increase in the FRET and BRET signals, highlighting its ability to affect the dimeric form of the receptor. Conclusions. Our results suggest that MT7 binds to a dimeric form of hM1 receptor, favouring the stability of this receptor state at the cellular level, probably by inducing some conformational rearrangements of the pre-existing muscarinic receptor homodimers.

Thermoplastic segmented polyurethanes (TPUs) are an important class of thermoplastic elastomers with a two-phase microstructure arising from the thermodynamic incompatibility between hard (HSs) and soft segments. This microphase separation observed on cooling from a homogeneous state is often combined with the solidification of either or both types of segments. In this study, the structuring mechanism of two TPUs with HSs based on 4,4'-diphenylmethane diisocyanate and 1,4-butanediol was investigated from rheological measurements. Hence, in addition to the structuring temperature influence, the effect of an applied preshear flow in the melt polymer was analyzed, in particular. The results clearly show an enhancement of the solidification kinetics by the preshear. Indeed, the measured structuring time can be reduced by more than 1 decade. Rheo-optical microscopy observations coupled with a shearing hot stage corroborated these results and showed the modification of the microstructure by the shear.

Nicotinic acetylcholine receptors (nAChR) are expressed on normal bronchial epithelial and nonsmall cell lung cancer (NSCLC) cells and are involved in cell growth regulation. Nicotine induced cell proliferation. The purpose of this study was to determine if interruption of autocrine nicotinic cholinergic signaling might inhibit A549 NSCLC cell growth. For this purpose alpha-Cobratoxin (alpha-CbT), a high affinity alpha7-nAChR antagonist was studied. Cell growth decrease was evaluated by Clonogenic and MTT assays. Evidence of apoptosis was identified staining cell with Annexin-V/PI. Characterization of the basal NF-kappaB activity was done using the Trans-AM NF-kappaB assay colorimetric kit. "In vivo" antitumour activity was evaluated in orthotopically transplanted nude mice monitored by In vivo Imaging System technology. alpha-CbT caused concentration-dependent cell growth decrease, mitochondrial apoptosis caspases-9 and 3-dependent, but caspase-2 and p53-independent and down-regulation of basal high levels of activated NF-kappaB. alpha-CbT treatment determines a significant reduction of tumor growth in nude mice orthotopically engrafted with A549-luciferase cells (4.6% of living cells vs. 31% in untreated mice). No sign of toxicity was reported related to treatment. These findings suggest that alpha7-nAChR antagonists namely alpha-CbT may be useful adjuvant for treatment of NSCLC and potentially other cancers.

Weak neurotoxins from snake venom are small proteins with five disulfide bonds, which have been shown to be poor binders of nicotinic acetylcholine receptors. We report on the cloning and sequencing of four cDNAs encoding weak neurotoxins from Naja sputatrix venom glands. The protein encoded by one of them, Wntx-5, has been synthesized by solid-phase synthesis and characterized. The physicochemical properties of the synthetic toxin (sWntx-5) agree with those anticipated for the natural toxin. We show that this toxin interacts with relatively low affinity (Kd = 180 nm) with the muscular-type acetylcholine receptor of the electric organ of T. marmorata, and with an even weaker affinity (90 µm) with the neuronal α7 receptor of chicken. Electrophysiological recordings using isolated mouse hemidiaphragm and frog cutaneous pectoris nerve–muscle preparations revealed no blocking activity of sWntx-5 at µm concentrations. Our data confirm previous observations that natural weak neurotoxins from cobras have poor affinity for nicotinic acetylcholine receptors.

The effect of neem kernel water extract (NKWE) in different concentrations on cassava mealybug, Phenacoccus manihoti, was investigated. A choice-test showed that neem treated cassava leaves were less attractive to first instar mealybugs than untreated leaves. Mealybugs that started feeding on treated leaves died in their second instar. A greenhouse experiment showed that three applications of NKWE at weekly intervals protected cassava against established early instar mealybugs. There were no significant differences between concentrations of 1, 10 and 25%. All concentrations were phytotoxic to a varying degree.

Synthetic approaches for new 2,5-disubstituted-1,3-thiazolidines are described. Steric and electronic effects of the N-substituent of the thiazolidine ring represent the major parameter in the rearrangement process. The nmr studies demonstrate that N-unsubstituted 2,5-disubstituted-1,3-thiazolidines exist as epimeric mixture, while the corresponding N-acetylated analogues exist as a conformer mixture.

Using a solid-phase assay, we found that 3H-labeled alpha Cobtx from Naja naja siamensis, a long-chain curaremimetic toxin, and 3H-labelled toxin alpha from Naja nigricollis, a short-chain toxin both bind specifically but with substantially different affinities (Kd = 4 x 10(-7) M and 50 x 10(-6) M) to fragment 185-199 (T alpha 185-199) of the alpha-subunit of the acetylcholine receptor (AcChoR) from Torpedo marmorata. Then we show that monoderivatizations of residues common to both long-chain and short-chain toxins (Tyr-25, Lys-27, Trp-29, and Lys-53) or to long-chain toxins only (Cys-30 and Cys-34) do not affect the binding of the toxins to T alpha 185-199, suggesting that none of these invariant residues in implicated in the recognition of this AcChoR region. alpha Cobtx and toxin alpha bind to the fragment 128-142 (T alpha 128-142) with more similar affinities (Kd = 3 x 10(-7) M and 1.4 x 10(-6) M) and their binding is dramatically affected by the single abolition of the positive charge of Lys-53, an invariant residue that contributes to AcChoR recognition. Therefore, the data indicate that Lys-53 more specifically recognizes the 128-142 region of AcChoR. Other monoderivatizations have no effect on toxin binding. The approach described in this paper may be of great help to identify toxin residues that establish direct contact with receptor fragments.

Synthetic approaches for new 2,5-disubstituted-1,3-thiazolidines are described. Steric and electronic effects of the N-substituent of the thiazolidine ring represent the major parameter in the rearrangement process. The nmr studies demonstrate that N-unsubstituted 2,5-disubstituted-1,3-thiazolidines exist as epimeric mixture, while the corresponding N-acetylated analogues exist as a conformer mixture.

The synthesis of new thiazolididone nucleoside analogues is described. Among the different proposed synthetic pathways, the condensation of various nucleic bases using TMSOTf and Et3N as coupling r...

Synthese, etudes structurales et evaluation antiretrovirale de 3'-aza-4'-thia-2',3'-didesoxynucleosides (analogues nucleosidiques de type thiazolidine). Considerant que des heterocycles mimant le cycle ribose pouvaient conferer a des analogues nucleosidiques des proprietes antivirales (HIV, HBV), un certain nombre de 3'-aza-4'-thia-2',3'-didesoxynucleosides ont ete synthetises. L'obtention de cette nouvelle classe d'analogues nucleosidiques a necessite la mise au point de synthese de 1,3-thiazolidine adequatement disubstituees en position 2 et 5. Le couplage des bases nucleiques sur les pseudo-sucres de type thiazolidine a ete realise a l'aide du chlorure d'etain(IV) comme catalyseur. Les deprotections donnant acces aux composes finaux 3'-aza-4'-thia-2',3'-didesoxynucleosides ont necessite l'utilisation du groupement protecteur N-fluorenylmethoxycarbonyl (N-Fmoc). Les groupements protecteurs N-acetyl et N-tosyl se sont averes inadequates lors de cette deprotection. Ces nouveaux composes nucleosidiques de type thiazolidine n'ont montre aucune activite antiretrovirale sur HIV.

On the basis of molecular modeling calculations using Gen Mol software, new 5-substituted-2′,3′-dideoxy-3′-thiauridine were designed as possible anti-HIV reverse transcriptase inhibitors. The synthesis of the key intermediate 5-carboxy-2′,3′-dideoxy-3′-thiauridine was achieved through the condensation of the fully silylated 5-carboxyuracil on 2-benzoyl methyl-5-acetoxy-1,3-oxathiolane using trimethylsilyl triflate (TMSOTf). This latter compound was condensed with 2-(N-tert-Butoxycarbonyl)-1 -aminoethane in the presence of N, N-diisopropylethylamine (DIEA). The subsequent carboxamide deprotection led to the final compounds. These new analogues were evaluated for their anti-HIV-1 activities on infected MT4 cells but no significant protection was observed. Electronic and structural parameters considered in this model were not sufficient to predict any active anti-HIV molecular structures.

The synthesis of new enantiomers of 5-o-carboranyl pyrimidine nucleosides is described. Some of these agents should be considered for boron neutron capture therapy.

A series of anti-HIV prodrugs possessing various polyaminated side arms have been developed. The incorporation of a N-Boc protected monoamine or diamine side arm into the backbone of the 2′,3′-dideoxy-3′-thiacytidine 1 (BCH-189) provided an increase in antiviral potency, which could be several orders magnitude greater than the parent drug (1) depending on the cell culture systems used (MT-4 or MDMs). Twenty six 2′,3′-dideoxy-3′-thiacytidine prodrugs which differ from each other by the length, the nature of the 5′-O function and the 5′-O or /and N-4 position on the nucleoside moiety were synthesized. Among this new series of prodrugs, several congeners (12c and 12a) were found to inhibit HIV-1 replication in cell culture with 50% effective concentrations ECso of 10 and 50 nM respectively, in MT-4 cells. Compound 12c was found more active on infected MDMs cells with 50% effective concentration of 0.01 nM. The synthesis and the antiviral properties of these compounds are discussed.