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The discovery of photonic crystals 30 years ago in conjunction with research advances in plasmonics and metamaterials, has inspired the concept of decameter scale metasurfaces, coined seismic metamaterials for an enhanced control of surface (Love and Rayleigh) and bulk (shear and pressure) elastodynamic waves. These powerful mathematical tools of coordinate transforms, effective medium and Floquet-Bloch theories which have revolutionized nanophotonics, can be translated in the language of civil engineering and geophysics. Experiments on seismic metamaterials made of buried elements in the soil demonstrate that the fore mentioned tools make a possible novel description of complex phenomena of soil-structure interaction during a seismic disturbance. But the concepts are already moving to more futuristic concepts and the same notions developed for structured soils are now used to examine the effects of buildings viewed as above surface resonators in megastructures such as metacities. But this perspective of future should not make us forget the heritage of the ancient peoples. Indeed, we finally point out the striking similarity between an invisible cloak design and the architecture of some ancient megastructures as the antique Gallo-Roman theaters and amphitheatres.

This paper analyzes the first checklist of insects from French Guiana. Compiled by a group of 70 experts based on published records, it comprises about 15 100 valid species names belonging to 20 orders and 322 families. Currently, about 17% of the species are only known from French Guiana or from the Guiana Shield region. Since Linné, the average rate of description has been 59 species per year, which has been increasing in the last 10 years. Based on a sample of recent taxonomic and faunistic papers covering 736 new species for French Guiana, 46% of the species came from new country records, the rest from new species descriptions. The rate of faunistic progress (new species or new records) is about 180 species per year over the last five years. Sixty-five percent of these faunal records came from non-professional entomologists and 74% of the holotypes of new species were collected by amateur entomologists. A rough extrapolation, using two different methods, provides a likely estimate of around 100000 species, the most conservative estimate being 67 000 species and the highest 184 000. Therefore, an estimated 80% of the species remain to be recorded and, in a best-case scenario, at least 270 years would be needed to complete the biotic inventory, at the current rate of species descriptions and distribution records. Although no order is exhaustively inventoried, the most in need of study are Diptera, Hymenoptera, Hemiptera and Trichoptera; and in absolute numbers, Coleoptera. These results and the fundamental role of non-professional entomologists in collecting and describing new species are discussed.

We outline some recent research advances on the control of elastic waves in thin and thick plates, that have occurred since the large scale experiment [S. Brûlé, Phys. Rev. Lett. 112, 133901 (2014)] that demonstrated significant interaction of surface seismic waves with holes structuring sedimentary soils at the meter scale. We further investigate the seismic wave trajectories of compressional body waves in soils structured with buildings. A significant substitution of soils by inclusions, acting as foundations, raises the question of the effective dynamic properties of these structured soils. Buildings, in the case of perfect elastic conditions for both soil and buildings, are shown to interact and strongly influence elastic body waves; such site-city seismic interactions were pointed out in [Guéguen et al., Bull. Seismol. Soc. Am. 92, 794–811 (2002)], and we investigate a variety of scenarios to illustrate the variety of behaviours possible.

We propose that wave propagation through a class of mechanical metamaterials opens unprecedented avenues in seismic wave protection based on spectral properties of auxetic-like metamaterials. The elastic parameters of these metamaterials like the bulk and shear moduli, the mass density, and even the Poisson ratio, can exhibit negative values in elastic stop bands. We show here that the propagation of seismic waves with frequencies ranging from 1 Hz to 40 Hz can be influenced by a decameter scale version of auxetic-like metamaterials buried in the soil, with the combined effects of impedance mismatch, local resonances and Bragg stop bands. More precisely, we numerically examine and illustrate the markedly different behaviors between the propagation of seismic waves through a homogeneous isotropic elastic medium (concrete) and an auxetic-like metamaterial plate consisting of 43 cells (40 m × 40 m × 40 m), utilized here as a foundation of a building one would like to protect from seismic site effects. This novel class of seismic metamaterials opens band gaps at frequencies compatible with seismic waves when they are designed appropriately, what makes them interesting candidates for seismic isolation structures.

Malaria symptoms are associated with the asexual multiplication of Plasmodium falciparum within human red blood cells (RBCs) and fever peaks coincide with the egress of daughter merozoites following the rupture of the parasitophorous vacuole (PV) and the RBC membranes. Over the last two decades, it has emerged that the release of competent merozoites is tightly regulated by a complex cascade of events, including the unusual multi-step activation mechanism of the pivotal subtilisin-like protease 1 (Sub1) that takes place in three different cellular compartments and remains poorly understood. Following an initial auto-maturation in the endoplasmic reticulum (ER) between its pro- and catalytic domains, the Sub1 prodomain (PD) undergoes further cleavages by the parasite aspartic protease plasmepsin X (PmX) within acidic secretory organelles that ultimately lead to full Sub1 activation upon discharge into the PV. Here, we report the crystal structure of full-length P. falciparum Sub1 (PfS1 FL ) and demonstrate, through structural, biochemical, and biophysical studies, that the atypical Plasmodium- specific Sub1 PD directly promotes the assembly of inactive enzyme homodimers at acidic pH, whereas Sub1 is primarily monomeric at neutral pH. Our results shed new light into the finely tuned Sub1 spatiotemporal activation during secretion, explaining how PmX processing and full activation of Sub1 can occur in different cellular compartments, and uncover a robust mechanism of pH-dependent subtilisin autoinhibition that plays a key role in P. falciparum merozoites egress from infected host cells.IMPORTANCEMalaria fever spikes are due to the rupture of infected erythrocytes, allowing the egress of Plasmodium sp. merozoites and further parasite propagation. This fleeting tightly regulated event involves a cascade of enzymes, culminating with the complex activation of the subtilisin-like protease 1, Sub1. Differently than other subtilisins, Sub1 activation strictly depends upon the processing by a parasite aspartic protease within acidic merozoite secretory organelles. However, Sub1 biological activity is required in the pH neutral parasitophorous vacuole, to prime effectors involved in the rupture of the vacuole and erythrocytic membranes. Here, we show that the unusual, parasite-specific Sub1 prodomain is directly responsible for its acidic-dependent dimerization and autoinhibition, required for protein secretion, before its full activation at neutral pH in a monomeric form. pH-dependent Sub1 dimerization defines a novel, essential regulatory element involved in the finely tuned spatiotemporal activation of the egress of competent Plasmodium merozoites., Competing Interests: The authors declare no conflict of interest.

Inosine 5'-monophosphate (IMP) dehydrogenase (IMPDH) is an ubiquitous enzyme that catalyzes the NAD + -dependent oxidation of inosine 5'-monophosphate into xanthosine 5'-monophosphate. This enzyme is formed of two distinct domains, a core domain where the catalytic reaction occurs, and a less-conserved Bateman domain. Our previous studies gave rise to the classification of bacterial IMPDHs into two classes, according to their oligomeric and kinetic properties. MgATP is a common effector but cause to different effects when it binds within the Bateman domain: it is either an allosteric activator for Class I IMPDHs or a modulator of the oligomeric state for Class II IMPDHs. To get insight into the role of the Bateman domain in the dissimilar properties of the two classes, deleted variants of the Bateman domain and chimeras issued from the interchange of the Bateman domain between the three selected IMPDHs have been generated and characterized using an integrative structural biology approach. Biochemical, biophysical, structural, and physiological studies of these variants unveil the Bateman domain as being the carrier of the molecular behaviors of both classes., (© 2023 The Protein Society.)

The human commensal fungus Candida albicans can attach to epithelia or indwelling medical devices and form biofilms, that are highly tolerant to antifungal drugs and can evade the immune response. The cell surface protein Pga59 has been shown to influence adhesion and biofilm formation. Here, we present evidence that Pga59 displays amyloid properties. Using electron microscopy, staining with an amyloid fibre-specific dye and X-ray diffraction experiments, we showed that the predicted amyloid-forming region of Pga59 is sufficient to build up an amyloid fibre in vitro and that recombinant Pga59 can also adopt a cross-β amyloid fibre architecture. Further, mutations impairing Pga59 amyloid assembly led to diminished adhesion to substrates and reduced biofilm production. Immunogold labelling on amyloid structures extracted from C. albicans revealed that Pga59 is used by the fungal cell to assemble amyloids within the cell wall in response to adhesion. Altogether, our results suggest that Pga59 amyloid properties are used by the fungal cell to mediate cell-substrate interactions and biofilm formation., (© 2023. The Author(s).)

The bacterial cell wall is a multi-layered mesh, whose major component is peptidoglycan (PG), a sugar polymer cross-linked by short peptide stems. During cell division, a careful balance of PG synthesis and degradation, precisely coordinated both in time and space, is necessary to prevent uncontrolled destruction of the cell wall. In Corynebacteriales, the D,L endopeptidase RipA has emerged as a major PG hydrolase for cell separation, and RipA defaults have major implications for virulence of the human pathogens Mycobacterium tuberculosis and Corynebacterium diphtheriae . However, the precise mechanisms by which RipA mediates cell separation remain elusive. Here we report phylogenetic, biochemical, and structural analysis of the Corynebacterium glutamicum homologue of RipA, Cg1735. The crystal structures of full-length Cg1735 in two different crystal forms revealed the C-terminal NlpC/P60 catalytic domain obtruded by its N-terminal conserved coiled-coil domain, which locks the enzyme in an autoinhibited state. We show that this autoinhibition is relieved by the extracellular core domain of the transmembrane septal protein Cg1604. The crystal structure of Cg1604 revealed a (β/α) protein with an overall topology similar to that of receiver domains from response regulator proteins. The atomic model of the Cg1735-Cg1604 complex, based on bioinformatical and mutational analysis, indicates that a conserved, distal-membrane helical insertion in Cg1604 is responsible for Cg1735 activation. The reported data provide important insights into how intracellular cell division signal(s), yet to be identified, control PG hydrolysis during RipA-mediated cell separation in Corynebacteriales .

The dengue virus nonstructural protein 1 (NS1) is a secreted virulence factor that modulates complement, activates immune cells and alters endothelial barriers. The molecular basis of these events remains incompletely understood. Here we describe a functional high affinity complex formed between NS1 and human high-density lipoproteins (HDL). Collapse of the soluble NS1 hexamer upon binding to the lipoprotein particle leads to the anchoring of amphipathic NS1 dimeric subunits into the HDL outer layer. The stable complex can be visualized by electron microscopy as a spherical HDL with rod-shaped NS1 dimers protruding from the surface. We further show that the assembly of NS1-HDL complexes triggers the production of pro-inflammatory cytokines in human primary macrophages while NS1 or HDL alone do not. Finally, we detect NS1 in complex with HDL and low-density lipoprotein (LDL) particles in the plasma of hospitalized dengue patients and observe NS1-apolipoprotein E-positive complexes accumulating overtime. The functional reprogramming of endogenous lipoprotein particles by NS1 as a means to exacerbate systemic inflammation during viral infection provides a new paradigm in dengue pathogenesis., (© 2022 The Authors.)

The C-terminus of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) protein E contains a PBM (PDZ-binding motif) targeting PDZ (PSD-95/Dlg/ZO-1) domains, which is identical to the PBM of SARS-CoV. The latter is involved in the pathogenicity of the virus. Recently, we identified 10 human PDZ-containing proteins showing significant interactions with SARS-CoV-2 protein E PBM. We selected several of them involved in cellular junctions and cell polarity (TJP1, PARD3, MLLT4, and LNX2) and MPP5/PALS1 previously shown to interact with SARS-CoV E PBM. Targeting cellular junctions and polarity components is a common strategy by viruses to hijack cell machinery to their advantage. In this study, we showed that these host PDZ domains TJP1, PARD3, MLLT4, LNX2, and MPP5/PALS1 interact in a PBM-dependent manner in vitro and colocalize with the full-length E protein in cellulo , sequestrating the PDZ domains to the Golgi compartment. We solved three crystal structures of complexes between human LNX2, MLLT4, and MPP5 PDZs and SARS-CoV-2 E PBM highlighting its binding preferences for several cellular targets. Finally, we showed different affinities for the PDZ domains with the original SARS-CoV-2 C-terminal sequence containing the PBM and the one of the beta variant that contains a mutation close to the PBM. The acquired mutations in the E protein localized near the PBM might have important effects both on the structure and the ion-channel activity of the E protein and on the host machinery targeted by the variants during the infection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Zhu, Alvarez, Wolff, Mechaly, Brûlé, Neitthoffer, Etienne-Manneville, Haouz, Boëda and Caillet-Saguy.)

The current COVID-19 pandemic illustrates the importance of obtaining reliable methods for the rapid detection of SARS-CoV-2. A highly specific and sensitive diagnostic test able to differentiate the SARS-CoV-2 virus from common human coronaviruses is therefore needed. Coronavirus nucleoprotein (N) localizes to the cytoplasm and the nucleolus and is required for viral RNA synthesis. N is the most abundant coronavirus protein, so it is of utmost importance to develop specific antibodies for its detection. In this study, we developed a sandwich immunoassay to recognize the SARS-CoV-2 N protein. We immunized one alpaca with recombinant SARS-CoV-2 N and constructed a large single variable domain on heavy chain (VHH) antibody library. After phage display selection, seven VHHs recognizing the full N protein were identified by ELISA. These VHHs did not recognize the nucleoproteins of the four common human coronaviruses. Hydrogen Deuterium eXchange-Mass Spectrometry (HDX-MS) analysis also showed that these VHHs mainly targeted conformational epitopes in either the C-terminal or the N-terminal domains. All VHHs were able to recognize SARS-CoV-2 in infected cells or on infected hamster tissues. Moreover, the VHHs could detect the SARS variants B.1.17/alpha, B.1.351/beta, and P1/gamma. We propose that this sandwich immunoassay could be applied to specifically detect the SARS-CoV-2 N in human nasal swabs., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Mycolactone is a diffusible lipid toxin produced by Mycobacterium ulcerans, the causative agent of Buruli ulcer disease. Altough bacterially derived mycolactone has been shown to traffic from cutaneous foci of infection to the bloodstream, the mechanisms underpinning its access to systemic circulation and import by host cells remain largely unknown. Using biophysical and cell-based approaches, we demonstrate that mycolactone specific association to serum albumin and lipoproteins is necessary for its solubilization and is a major mechanism to regulate its bioavailability. We also demonstrate that Scavenger Receptor (SR)-B1 contributes to the cellular uptake of mycolactone. Overall, we suggest a new mechanism of transport and cell entry, challenging the dogma that the toxin enters host cells via passive diffusion., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Tello Rubio, Bugault, Baudon, Raynal, Brûlé, Morel, Saint-Auret, Blanchard, Demangel and Guenin-Macé.)

Intrinsic viscosity is a key hydrodynamic parameter to understand molecular structure and hydration, as well as intramolecular interactions. Commercially available instruments measure intrinsic viscosity by recording the macromolecular mobility in a capillary. These instruments monitor Taylor dispersion using an absorbance or fluorescence detector. By design, these instruments behave like U-tube viscometers. To our knowledge, there are no studies to date showing that the Viscosizer TD instrument (Malvern-Panalytical) is able to measure the intrinsic viscosity of macromolecules. In this study, we then performed our assays on the Poly(ethylene oxide) polymer (PEO), used classically as a standard for viscometry measurements and on three model proteins: the bovine serum albumin (BSA), the bevacizumab monoclonal antibody, and the RTX Repeat Domain (RD) of the adenylate cyclase toxin of Bordetella pertussis (CyaA). The presence of P20 in the samples is critical to get reliable results. The data obtained with our in-house protocol show a strong correlation with intrinsic viscosity values obtained using conventional techniques. However, with respect to them, our measurements could be performed at relatively low concentrations, between 2 and 5 mg/ml, using only 7 µL per injection. Altogether, our results show that the Viscosizer TD instrument is able to measure intrinsic viscosities in a straightforward manner. This simple and innovative approach should give a new boost to intrinsic viscosity measurements and should reignite the interest of biophysicists, immunologists, structural biologists and other researchers for this key physicochemical parameter.

To stop the COVID-19 pandemic due to the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), which caused more than 2.5 million deaths to date, new antiviral molecules are urgently needed. The replication of SARS-CoV-2 requires the RNA-dependent RNA polymerase (RdRp), making RdRp an excellent target for antiviral agents. RdRp is a multi-subunit complex composed of 3 viral proteins named nsp7, nsp8 and nsp12 that ensure the ~30 kb RNA genome's transcription and replication. The main strategies employed so far for the overproduction of RdRp consist of expressing and purifying the three subunits separately before assembling the complex in vitro. However, nsp12 shows limited solubility in bacterial expression systems and is often produced in insect cells. Here, we describe an alternative strategy to co-express the full SARS-CoV-2 RdRp in E. coli, using a single plasmid. Characterization of the purified recombinant SARS-CoV-2 RdRp shows that it forms a complex with the expected (nsp7)(nsp8)2(nsp12) stoichiometry. RNA polymerization activity was measured using primer-extension assays showing that the purified enzyme is functional. The purification protocol can be achieved in one single day, surpassing in speed all other published protocols. Our construct is ideally suited for screening RdRp and its variants against very large chemical compounds libraries and has been made available to the scientific community through the Addgene plasmid depository (Addgene ID: 165451)., Competing Interests: The authors have declared that no competing interests exist.

The molecular mechanisms and forces involved in the translocation of bacterial toxins into host cells are still a matter of intense research. The adenylate cyclase (CyaA) toxin from Bordetella pertussis displays a unique intoxication pathway in which its catalytic domain is directly translocated across target cell membranes. The CyaA translocation region contains a segment, P454 (residues 454-484), which exhibits membrane-active properties related to antimicrobial peptides. Herein, the results show that this peptide is able to translocate across membranes and to interact with calmodulin (CaM). Structural and biophysical analyses reveal the key residues of P454 involved in membrane destabilization and calmodulin binding. Mutational analysis demonstrates that these residues play a crucial role in CyaA translocation into target cells. In addition, calmidazolium, a calmodulin inhibitor, efficiently blocks CyaA internalization. It is proposed that after CyaA binding to target cells, the P454 segment destabilizes the plasma membrane, translocates across the lipid bilayer and binds calmodulin. Trapping of CyaA by the CaM:P454 interaction in the cytosol may assist the entry of the N-terminal catalytic domain by converting the stochastic motion of the polypeptide chain through the membrane into an efficient vectorial chain translocation into host cells., Competing Interests: The authors declare no conflict of interest., (© 2021 The Authors. Advanced Science published by Wiley‐VCH GmbH.)

PDZ domains are small globular domains involved in protein-protein interactions. They participate in a wide range of critical cellular processes. These domains, very abundant in the human proteome, are widely studied by high-throughput interactomics approaches and by biophysical and structural methods. However, the quality of the results is strongly related to the optimal folding and solubility of the domains. We provide here a detailed description of protocols for a strict quality assessment of the PDZ constructs. We describe appropriate experimental approaches that have been selected to overcome the small size of such domains to check the purity, identity, homogeneity, stability, and folding of samples.

One essential prerequisite of any experiment involving a purified protein, such as interaction studies or structural and biophysical characterization, is to work with a "good-quality" sample in order to ensure reproducibility and reliability of the data. Here, we define a "good-quality" sample as a protein preparation that fulfills three criteria: (1) the preparation contains a protein that is pure and soluble and exhibits structural and functional integrity, (2) the protein must be structurally homogeneous, and (3) the preparation must be reproducible. To ensure effective quality control (QC) of all these parameters, we suggest to follow a simple workflow involving the use of gel electrophoresis, light scattering, and spectroscopic experiments. We describe the techniques used in every step of this workflow and provide easy-to-use standard protocols for each step.

Hearing is a mechanical and neurochemical process, which occurs in the hair cells of inner ear that converts the sound vibrations into electrical signals transmitted to the brain. The multi-PDZ scaffolding protein whirlin plays a critical role in the formation and function of stereocilia exposed at the surface of hair cells. In this article, we reported seven stereociliary proteins that encode PDZ binding motifs (PBM) and interact with whirlin PDZ3, where four of them are first reported. We solved the atomic resolution structures of complexes between whirlin PDZ3 and the PBMs of myosin 15a, CASK, harmonin a1 and taperin. Interestingly, the PBM of CASK and taperin are rare non-canonical PBM, which are not localized at the extreme C terminus. This large capacity to accommodate various partners could be related to the distinct functions of whirlin at different stages of the hair cell development., (Copyright © 2020. Published by Elsevier Ltd.)

Omalizumab is an anti-IgE monoclonal antibody (mAb) approved for the treatment of severe asthma and chronic spontaneous urticaria. Use of omalizumab is associated with reported side effects ranging from local skin inflammation at the injection site to systemic anaphylaxis. To date, the mechanisms through which omalizumab induces adverse reactions are still unknown. Here, we demonstrated that immune complexes formed between omalizumab and IgE can induce both skin inflammation and anaphylaxis through engagement of IgG receptors (FcγRs) in FcγR-humanized mice. We further developed an Fc-engineered mutant version of omalizumab, and demonstrated that this mAb is equally potent as omalizumab at blocking IgE-mediated allergic reactions, but does not induce FcγR-dependent adverse reactions. Overall, our data indicate that omalizumab can induce skin inflammation and anaphylaxis by engaging FcγRs, and demonstrate that Fc-engineered versions of the mAb could be used to reduce such adverse reactions.

Ticks are the most important vectors of pathogens affecting both domestic and wild animals worldwide. Hard tick feeding is a slow process-taking up to several days-and necessitates extended control over the host response. The success of the feeding process depends upon injection of tick saliva, which not only controls host hemostasis and wound healing, but also subverts the host immune response to avoid tick rejection that creates a favorable niche for the survival and propagation of diverse tick-borne pathogens. Here, we report on the molecular and biochemical features and functions of an Ixodes ricinus serine protease inhibitor (IrSPI). We characterize IrSPI as a Kunitz elastase inhibitor that is overexpressed in several tick organs-especially salivary glands-during blood-feeding. We also demonstrated that when IrSPI is injected into the host through saliva, it had no impact on tissue factor pathway-induced coagulation, fibrinolysis, endothelial cell angiogenesis or apoptosis, but the protein exhibits immunomodulatory activity. In particular, IrSPI represses proliferation of CD4 + T lymphocytes and proinflammatory cytokine secretion from both splenocytes and macrophages. Our study contributes valuable knowledge to tick-host interactions and provides insights that could be further exploited to design anti-tick vaccines targeting this immunomodulator implicated in I. ricinus feeding.

Protozoan pathogens secrete nanosized particles called extracellular vesicles (EVs) to facilitate their survival and chronic infection. Here, we show the inhibition by Plasmodium berghei NK65 blood stage-derived EVs of the proliferative response of CD4 + T cells in response to antigen presentation. Importantly, these results were confirmed in vivo by the capacity of EVs to diminish the ovalbumin-specific delayed type hypersensitivity response. We identified two proteins associated with EVs, the histamine releasing factor (HRF) and the elongation factor 1α (EF-1α) that were found to have immunosuppressive activities. Interestingly, in contrast to WT parasites, EVs from genetically HRF- and EF-1α-deficient parasites failed to inhibit T cell responses in vitro and in vivo. At the level of T cells, we demonstrated that EVs from WT parasites dephosphorylate key molecules (PLCγ1, Akt, and ERK) of the T cell receptor signalling cascade. Remarkably, immunisation with EF-1α alone or in combination with HRF conferred a long-lasting antiparasite protection and immune memory. In conclusion, we identified a new mechanism by which P. berghei-derived EVs exert their immunosuppressive functions by altering T cell responses. The identification of two highly conserved immune suppressive factors offers new conceptual strategies to overcome EV-mediated immune suppression in malaria-infected individuals., (© 2019 John Wiley & Sons Ltd.)

As all the viruses belonging to the Mononegavirales order, the nonsegmented negative-strand RNA genome of respiratory syncytial virus (RSV) is encapsidated by the viral nucleoprotein N. N protein polymerizes along the genomic and anti-genomic RNAs during replication. This requires the maintenance of the neosynthesized N protein in a monomeric and RNA-free form by the viral phosphoprotein P that plays the role of a chaperone protein, forming a soluble N 0 -P complex. We have previously demonstrated that residues 1-30 of P specifically bind to N 0 Here, to isolate a stable N 0 -P complex suitable for structural studies, we used the N-terminal peptide of P (P40) to purify truncated forms of the N protein. We show that to purify a stable N 0 -P-like complex, a deletion of the first 30 N-terminal residues of N (N Δ30 ) is required to impair N oligomerization, whereas the presence of a full-length C-arm of N is required to inhibit RNA binding. We generated structural models of the RSV N 0 -P with biophysical approaches, including hydrodynamic measurements and small-angle X-ray scattering (SAXS), coupled with biochemical and functional analyses of human RSV (hRSV) N Δ30 mutants. These models suggest a strong structural homology between the hRSV and the human metapneumovirus (hMPV) N 0 -P complexes. In both complexes, the P40-binding sites on N 0 appear to be similar, and the C-arm of N provides a high flexibility and a propensity to interact with the N RNA groove. These findings reveal two potential sites to target on N 0 -P for the development of RSV antivirals., (© 2019 Esneau et al.)

Pathogenic Leptospira bacteria are the causative agents of leptospirosis, a zoonotic disease affecting animals and humans worldwide. These pathogenic species have the ability to rapidly cross host tissue barriers by a yet unknown mechanism. A comparative analysis of pathogens and saprophytes revealed a higher abundance of genes encoding proteins with leucine-rich repeat (LRR) domains in the genomes of pathogens. In other bacterial pathogens, proteins with LRR domains have been shown to be involved in mediating host cell attachment and invasion. One protein from the pathogenic species Leptospira interrogans, LIC10831, has been previously analysed via X-ray crystallography, with findings suggesting it may be an important bacterial adhesin. Herein we show that LIC10831 elicits an antibody response in infected animals, is actively secreted by the bacterium, and binds human E- and VE-cadherins. These results provide biochemical and cellular evidences of LRR protein-mediated host-pathogen interactions and identify a new multireceptor binding protein from this infectious Leptospira species., (© 2018 John Wiley & Sons Ltd.)

PolD is an archaeal replicative DNA polymerase (DNAP) made of a proofreading exonuclease subunit (DP1) and a larger polymerase catalytic subunit (DP2). Recently, we reported the individual crystal structures of the DP1 and DP2 catalytic cores, thereby revealing that PolD is an atypical DNAP that has all functional properties of a replicative DNAP but with the catalytic core of an RNA polymerase (RNAP). We now report the DNA-bound cryo-electron microscopy (cryo-EM) structure of the heterodimeric DP1-DP2 PolD complex from Pyrococcus abyssi, revealing a unique DNA-binding site. Comparison of PolD and RNAPs extends their structural similarities and brings to light the minimal catalytic core shared by all cellular transcriptases. Finally, elucidating the structure of the PolD DP1-DP2 interface, which is conserved in all eukaryotic replicative DNAPs, clarifies their evolutionary relationships with PolD and sheds light on the domain acquisition and exchange mechanism that occurred during the evolution of the eukaryotic replisome., Competing Interests: The authors have declared that no competing interests exist.

We show that seismic energy simulated by an artificial source that mainly propagates Rayleigh surface waves, is focused in structured soil made of a grid of holes distributed in the ground. We carry out large-scale field tests with a structured soil made of a grid consisting of cylindrical and vertical holes in the ground and a low frequency artificial source (<10 Hz). This allows the identification of a distribution of energy inside the grid, which can be interpreted as the consequence of a dynamic anisotropy akin to an effective negative refraction index. Such a flat lens reminiscent of what Veselago and Pendry envisioned for light opens avenues in seismic metamaterials to counteract partially or totally the most devastating components of seismic signals.

We begin with a brief historical survey of discoveries of quasi-crystals and graphene, and then introduce the concept of transformation crystallography, which consists of the application of geometric transforms to periodic structures. We consider motifs with three-fold, four-fold and six-fold symmetries according to the crystallographic restriction theorem. Furthermore, we define motifs with five-fold symmetry such as quasi-crystals generated by a cut-and-projection method from periodic structures in higher-dimensional space. We analyze elastic wave propagation in the transformed crystals and (Penrose-type) quasi-crystals with the finite difference time domain freeware SimSonic. We consider geometric transforms underpinning the design of seismic cloaks with square, circular, elliptical and peanut shapes in the context of honeycomb crystals that can be viewed as scaled-up versions of graphene. Interestingly, the use of morphing techniques leads to the design of cloaks with interpolated geometries reminiscent of Victor Vasarely's artwork. Employing the case of transformed graphene-like (honeycomb) structures allows one to draw useful analogies between large-scale seismic metamaterials such as soils structured with columns of concrete or grout with soil and nanoscale biochemical metamaterials. We further identify similarities in designs of cloaks for elastodynamic and hydrodynamic waves and cloaks for diffusion (heat or mass) processes, as these are underpinned by geometric transforms. Experimental data extracted from field test analysis of soil structured with boreholes demonstrates the application of crystallography to large scale phononic crystals, coined as seismic metamaterials, as they might exhibit low frequency stop bands. This brings us to the outlook of mechanical metamaterials, with control of phonon emission in graphene through extreme anisotropy, attenuation of vibrations of suspension bridges via low frequency stop bands and the concept of transformed meta-cities. We conclude that these novel materials hold strong applications spanning different disciplines or across different scales from biophysics to geophysics.

Evidence points to a role of the mammalian target of rapamycin (mTOR) signaling pathway as a regulator of adiposity, yet its involvement as a mediator of the positive actions of peroxisome proliferator-activated receptor (PPAR)γ agonism on lipemia, fat accretion, lipid uptake, and its major determinant lipoprotein lipase (LPL) remains to be elucidated. Herein we evaluated the plasma lipid profile, triacylglycerol (TAG) secretion rates, and adipose tissue LPL-dependent lipid uptake, LPL expression/activity, and expression profile of other lipid metabolism genes in rats treated with the PPARγ agonist rosiglitazone (15 mg/kg/day) in combination or not with the mTOR inhibitor rapamycin (2 mg/kg/day) for 15 days. Rosiglitazone stimulated adipose tissue mTOR complex 1 and AMPK and induced TAG-derived lipid uptake (136%), LPL mRNA/activity (2- to 6-fold), and fat accretion in subcutaneous (but not visceral) white adipose tissue (WAT; 50%) and in brown adipose tissue (BAT; 266%). Chronic mTOR inhibition attenuated the upregulation of lipid uptake, LPL expression/activity, and fat accretion induced by PPARγ activation in both subcutaneous WAT and BAT, which resulted in hyperlipidemia. In contrast, rapamycin did not affect most of the other WAT lipogenic genes upregulated by rosiglitazone. Together these findings demonstrate that mTOR is a major regulator of adipose tissue LPL-mediated lipid uptake and a critical mediator of the hypolipidemic and lipogenic actions of PPARγ activation.

Objective: The mammalian target of rapamycin (mTOR)/p70 S6 kinase 1 (S6K1) pathway is a critical signaling component in the development of obesity-linked insulin resistance and operates a nutrient-sensing negative feedback loop toward the phosphatidylinositol 3-kinase (PI 3-kinase)/Akt pathway. Whereas acute treatment of insulin target cells with the mTOR complex 1 (mTORC1) inhibitor rapamycin prevents nutrient-induced insulin resistance, the chronic effect of rapamycin on insulin sensitivity and glucose metabolism in vivo remains elusive., Research Design and Methods: To assess the metabolic effects of chronic inhibition of the mTORC1/S6K1 pathway, rats were treated with rapamycin (2 mg/kg/day) or vehicle for 15 days before metabolic phenotyping., Results: Chronic rapamycin treatment reduced adiposity and fat cell number, which was associated with a coordinated downregulation of genes involved in both lipid uptake and output. Rapamycin treatment also promoted insulin resistance, severe glucose intolerance, and increased gluconeogenesis. The latter was associated with elevated expression of hepatic gluconeogenic master genes, PEPCK and G6Pase, and increased expression of the transcriptional coactivator peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) as well as enhanced nuclear recruitment of FoxO1, CRTC2, and CREB. These changes were observed despite normal activation of the insulin receptor substrate/PI 3-kinase/Akt axis in liver of rapamycin-treated rats, as expected from the blockade of the mTORC1/S6K1 negative feedback loop., Conclusions: These findings unravel a novel mechanism by which mTORC1/S6K1 controls gluconeogenesis through modulation of several key transcriptional factors. The robust induction of the gluconeogenic program in liver of rapamycin-treated rats underlies the development of severe glucose intolerance even in the face of preserved hepatic insulin signaling to Akt and despite a modest reduction in adiposity.

Dysfunctional mTORC1 signaling is associated with a number of human pathologies owing to its central role in controlling cell growth, proliferation, and metabolism. Regulation of mTORC1 is achieved by the integration of multiple inputs, including those of mitogens, nutrients, and energy. It is thought that agents that increase the cellular AMP/ATP ratio, such as the antidiabetic biguanides metformin and phenformin, inhibit mTORC1 through AMPK activation of TSC1/2-dependent or -independent mechanisms. Unexpectedly, we found that biguanides inhibit mTORC1 signaling, not only in the absence of TSC1/2 but also in the absence of AMPK. Consistent with these observations, in two distinct preclinical models of cancer and diabetes, metformin acts to suppress mTORC1 signaling in an AMPK-independent manner. We found that the ability of biguanides to inhibit mTORC1 activation and signaling is, instead, dependent on the Rag GTPases.

Endocrine effects of adipose-derived adiponectin on skeletal muscle have been shown to account, at least in part, for the anti-diabetic effects of this adipokine. Recently, the concept of myokines has gained credence, and the potential for skeletal muscle to produce adiponectin has been suggested. Here we demonstrated an increased level of adiponectin mRNA and protein expression as well as protein secretion in response to rosiglitazone treatment in L6 muscle cells. This correlated with the ability of rosiglitazone to enhance insulin sensitivity for stimulation of protein kinase B (Akt) phosphorylation and glucose transport; rosiglitazone also corrected high-glucose-induced insulin resistance in L6 cells. Overexpression of adiponectin confirmed the functional significance of local production of adiponectin in muscle cells via elevated glucose uptake and increased insulin sensitivity. In obese diabetic db/db mice, there was a change in the adiponectin expression profile in soleus and extensor digitorum longus (EDL) muscle with less high molecular weight (HMW) and more medium (MMW)/low (LMW) molecular weight species detected. Induction of obesity and insulin resistance in rats by feeding a high-fat high-sucrose diet also led to decreased muscle HMW adiponectin content that could be corrected by rosiglitazone treatment. In summary, we show the ability of skeletal muscle cells to produce adiponectin, which can mediate autocrine metabolic effects, thus establishing adiponectin as a bona fide myokine. We also demonstrate that skeletal muscle adiponectin production is altered in animal models of obesity and diabetes and that these changes can be corrected by rosiglitazone.

We investigated cardiac hypertrophy elicited by rosiglitazone treatment at the level of protein synthesis/degradation, mTOR, MAPK and AMPK signalling pathways, cardiac function and aspects of carbohydrate/lipid metabolism. Hearts of rats treated or not with rosiglitazone (15 mg/kg day) for 21 days were evaluated for gene expression, protein synthesis, proteasome and calpain activities, signalling pathways, and function by echocardiography. Rosiglitazone induced eccentric heart hypertrophy associated with increased expression of ANP, BNP, collagen I and III and fibronectin, reduced heart rate and increased stroke volume. Rosiglitazone robustly increased heart glycogen content ( approximately 400%), an effect associated with increases in glycogenin and UDPG-PPL mRNA levels and glucose uptake, and a reduction in glycogen phosphorylase expression and activity. Cardiac triglyceride content, lipoprotein lipase activity and mRNA levels of enzymes involved in fatty acid oxidation were also reduced by the agonist. Rosiglitazone-induced cardiac hypertrophy was associated with an increase in myofibrillar protein content and turnover (increased synthesis and an enhancement of calpain-mediated myofibrillar degradation). In contrast, 26S beta5 chymotryptic proteasome activity and mRNA levels of 20S beta2 and beta5 and 19S RPN 2 proteasome subunits along with the ubiquitin ligases atrogin and CHIP were all reduced by rosiglitazone. These morphological and biochemical changes were associated with marked activation of the key growth-promoting mTOR signalling pathway, whose pharmacological inhibition with rapamycin completely blocked cardiac hypertrophy induced by rosiglitazone. The study demonstrates that both arms of protein balance are involved in rosiglitazone-induced cardiac hypertrophy, and establishes the mTOR pathway as a novel important mediator therein.

S6K1 has emerged as a critical signaling component in the development of insulin resistance through phosphorylation and inhibition of IRS-1 function. This effect can be triggered directly by nutrients such as amino acids or by insulin through a homeostatic negative-feedback loop. However, the role of S6K1 in mediating IRS-1 phosphorylation in a physiological setting of nutrient overload is unresolved. Here we show that S6K1 directly phosphorylates IRS-1 Ser-1101 in vitro in the C-terminal domain of the protein and that mutation of this site largely blocks the ability of amino acids to suppress IRS-1 tyrosine and Akt phosphorylation. Consistent with this finding, phosphorylation of IRS-1 Ser-1101 is increased in the liver of obese db/db and wild-type, but not S6K1(-/-), mice maintained on a high-fat diet and is blocked by siRNA knockdown of S6K1 protein. Finally, infusion of amino acids in humans leads to the concomitant activation of S6K1, phosphorylation of IRS-1 Ser-1101, a reduction in IRS-1 function, and insulin resistance in skeletal muscle. These findings indicate that nutrient- and hormonal-dependent activation of S6K1 causes insulin resistance in mice and humans, in part, by mediating IRS-1 Ser-1101 phosphorylation.