The development of resistance to an inhibitor of a cellular protein reveals a critical interaction between the enterovirus protein 2C and a small GTPase Arf1.

The cellular protein GBF1, an activator of Arf GTPases (ArfGEF: Arf guanine nucleotide exchange factor), is recruited to the replication organelles of enteroviruses through interaction with the viral protein 3A, and its ArfGEF activity is required for viral replication, however how GBF1-dependent Arf activation supports the infection remains enigmatic. Here, we investigated the development of resistance of poliovirus, a prototype enterovirus, to increasing concentrations of brefeldin A (BFA), an inhibitor of GBF1. High level of resistance required a gradual accumulation of multiple mutations in the viral protein 2C. The 2C mutations conferred BFA resistance even in the context of a 3A mutant previously shown to be defective in the recruitment of GBF1 to replication organelles, and in cells depleted of GBF1, suggesting a GBF1-independent replication mechanism. Still, activated Arfs accumulated on the replication organelles of this mutant even in the presence of BFA, its replication was inhibited by a pan-ArfGEF inhibitor LM11, and the BFA-resistant phenotype was compromised in Arf1-knockout cells. Importantly, the mutations strongly increased the interaction of 2C with the activated form of Arf1. Analysis of other enteroviruses revealed a particularly strong interaction of 2C of human rhinovirus 1A with activated Arf1. Accordingly, the replication of this virus was significantly less sensitive to BFA than that of poliovirus. Thus, our data demonstrate that enterovirus 2Cs may behave like Arf1 effector proteins and that GBF1 but not Arf activation can be dispensable for enterovirus replication. These findings have important implications for the development of host-targeted anti-viral therapeutics.

A Proximity biotinylation assay with a host protein bait reveals multiple factors modulating enterovirus replication.

As ultimate parasites, viruses depend on host factors for every step of their life cycle. On the other hand, cells evolved multiple mechanisms of detecting and interfering with viral replication. Yet, our understanding of the complex ensembles of pro- and anti-viral factors is very limited in virtually every virus-cell system. Here we investigated the proteins recruited to the replication organelles of poliovirus, a representative of the genus Enterovirus of the Picornaviridae family. We took advantage of a strict dependence of enterovirus replication on a host protein GBF1, and established a stable cell line expressing a truncated GBF1 fused to APEX2 peroxidase that effectively supported viral replication upon inhibition of the endogenous GBF1. This construct biotinylated multiple host and viral proteins on the replication organelles. Among the viral proteins, the polyprotein cleavage intermediates were overrepresented, suggesting that the GBF1 environment is linked to viral polyprotein processing. The proteomics characterization of biotinylated host proteins identified multiple proteins previously associated with enterovirus replication, as well as more than 200 new factors recruited to the replication organelles. RNA metabolism proteins, many of which normally localize in the nucleus, constituted the largest group, underscoring the massive release of nuclear factors into the cytoplasm of infected cells and their involvement in viral replication. Functional analysis of several newly identified proteins revealed both pro- and anti-viral factors, including a novel component of infection-induced stress granules. Depletion of these proteins similarly affected the replication of diverse enteroviruses indicating broad conservation of the replication mechanisms. Thus, our data significantly expand the knowledge of the composition of enterovirus replication organelles, provide new insights into viral replication, and offer a novel resource for identifying targets for anti-viral interventions.

Chemical bonding effects in Sc compounds studied using X-ray absorption and X-ray photoelectron spectroscopies.

Advances on understanding the nature of the chemical bonding and electron correlation effects during the X-ray absorption process in ionic-covalent metal complexes has been achieved for most of the transition elements, except for scandium, due to the lack of a systematic series of spectroscopic reference spectra and the shortage of standard crystallographic data on scandium compounds. To close the gap, the chemical bonding effects in eight Sc compounds are studied using X-ray absorption spectroscopy (XAS) at Sc K and L2,3 absorption edges and X-ray photoelectron spectroscopy (XPS). Indeed, the fine structure of the XAS Sc K edge reflects the chemical sp3-like bond formed between scandium and the ligand while the L2,3 edge and the pre-edge features of the K-edge provide a direct insight into the crystal field parameters at the Sc site in the coordination compound. The XPS data provide the information on binding energies of the core electrons involved in the electron transitions caused by the absorption of high energy X-rays. XAS and XPS complement each other by accessing the information on Sc structure on bulk and the surface. Herein, comprehensive information on the electronic structure of well-known crystalline materials based on Sc is given with spectroscopic fingerprints X-ray data. This will help to predict the formation of chemical bonds in the unknown components via the systematic evaluation of the available spectroscopic fingerprints.

Unlocking a Dual-Channel Pathway in CO2 Hydrogenation to Methanol over Single-Site Zirconium on Amorphous Silica.

Converting CO2 into methanol on a large scale is of great significance in the sustainable methanol economy. Zirconia species are considered to be an essential support in Cu-based catalysts due to their excellent properties for CO2 adsorption and activation. However, the evolution of Zr species during the reaction and the effect of their structure on the reaction pathways remain unclear. Herein, single-site Zr species in an amorphous SiO2 matrix are created by enhancing the Zr-Si interaction in Cu/ZrO2 -SiO2 catalysts. In situ X-ray absorption spectroscopy (XAS) reveals that the coordination environment of single-site Zr is sensitive to the atmosphere and reaction conditions. We demonstrate that the CO2 adsorption occurs preferably on the interface of Cu and single-site Zr rather than on ZrO2 nanoparticles. Methanol synthesis in reverse water-gas-shift (RWGS)+CO-hydro pathway is verified only over single-dispersed Zr sites, whereas the ordinary formate pathway occurs on ZrO2 nanoparticles. Thus, it expands a non-competitive parallel pathway as a supplement to the dominant formate pathway, resulting in the enhancement of Cu activity sixfold and twofold based on Cu/SiO2 and Cu/ZrO2 catalysts, respectively. The establishment of this dual-channel pathway by single-site Zr species in this work opens new horizons for understanding the role of atomically dispersed oxides in catalysis science.

Fundamental Structural and Electronic Understanding of Palladium Catalysts on Nitride and Oxide Supports.

The nature of the support can fundamentally affect the function of a heterogeneous catalyst. For the novel type of isolated metal atom catalysts, sometimes referred to as single-atom catalysts, systematic correlations are still rare. Here, we report a general finding that Pd on nitride supports (non-metal and metal nitride) features a higher oxidation state compared to that on oxide supports (non-metal and metal oxide). Through thorough oxidation state investigations by X-ray absorption spectroscopy (XAS), X-ray photoelectron spectroscopy (XPS), CO-DRIFTS, and density functional theory (DFT) coupled with Bader charge analysis, it is found that Pd atoms prefer to interact with surface hydroxyl group to form a Pd(OH)x species on oxide supports, while on nitride supports, Pd atoms incorporate into the surface structure in the form of Pd-N bonds. Moreover, a correlation was built between the formal oxidation state and computational Bader charge, based on the periodic trend in electronegativity.

Probing the Nature of Zinc in Copper-Zinc-Zirconium Catalysts by Operando Spectroscopies for CO2 Hydrogenation to Methanol.

Active Zn species in Cu-based methanol synthesis catalysts have not been clearly identified yet due to their complex nature and dynamic structural changes during reactions. Herein, atomically dispersed Zn on ZrO2 support is established in Cu-based catalysts by separating Zn and Zr components from Cu (Cu-ZnZr) via the double-nozzle flame spray pyrolysis (DFSP) method. It exhibits superiority in methanol selectivity and yield compared to those with Cu-ZnO interface and isolated ZnO nanoparticles. Operando X-ray absorption spectroscopy (XAS) reveals that the atomically dispersed Zn species are induced during the reaction due to the strengthened Zn-Zr interaction. They can suppress formate decomposition to CO and decrease the H2 dissociation energy, shifting the reaction to methanol production. This work enlightens the rational design of unique Zn species by regulating coordination environments and offers a new perspective for exploring complex interactions in multi-component catalysts.

In Situ Investigations on Structural Evolutions during the Facile Synthesis of Cubic α-MoC1-x Catalysts.

Cubic α-phase molybdenum carbides (α-MoC1-x) exhibit great potential in hydrogen production at low temperatures due to their excellent activity in water dissociation. However, the design strategies of α-MoC1-x are severely restricted by the harsh synthesis conditions, which involve multistep ammonification and carburization or the utilization of a significant amount of noble metals. Herein, high-purity α-MoC1-x synthesis in a one-step carburization process was achieved with the assistance of a trace amount of Rh (0.02%). The structural evolution of Mo species during phase transition was monitored via qualitative and quantitative analysis by in situ X-ray diffraction (XRD) and in situ X-ray absorption spectroscopy (XAS), respectively. Environmental transmission electron microscopy (ETEM) was used to follow the visual changes. We reveal that the reduction of monoclinic MoO3 to cubic oxygen-deficient Mo oxide (MoOx) at low temperatures owing to the promoted H2 activation on Rh sites is vital to the following carbon atom insertion and transformation to α-MoC1-x, making the carburization follow the topological route. The systematic analysis of the relationship between the reduction behavior and the structural evolution supplies a feasible strategy for the α-MoC1-x synthesis, and in situ characterizations shed light on controlling the phase transformation during carburization.

Hybrid Implants Based on Calcium-Magnesium Silicate Ceramics Diopside as a Carrier of Recombinant BMP-2 and Demineralized Bone Matrix as a Scaffold: Dynamics of Reparative Osteogenesis in a Mouse Craniotomy Model.

Calcium-magnesium silicate ceramics, diopside, is a promising material for use in bone plastics, but until now the possibility of its use as a carrier of recombinant bone morphogenetic protein-2 (BMP-2) has not been studied, as well as the features of reparative osteogenesis mediated by the materials based on diopside with BMP-2. Powder of calcium-magnesium silicate ceramics was obtained by solid-state synthesis using biowaste - rice husks and egg shells - as source components. Main phase of the obtained ceramics was diopside. The obtained particles were irregularly shaped with an average size of about 2.3 µm and ~20% porosity; average pore size was about 24 nm, which allowed the material to be classified as mesoporous. Diopside powder adsorbs more than 150 µg of recombinant BMP-2 per milligram, which exceeds binding capacity of hydroxyapatite, a calcium-phosphate ceramic often used in hybrid implants, by more than 3 times. In vitro release kinetics of BMP-2 was characterized by a burst release in the first 2 days and a sustained release of approximately 0.4 to 0.5% of the loaded protein over the following 7 days. In vivo experiments were performed with a mouse model of cranial defects of critical size with implantation of a suspension of diopside powder with/without BMP-2 in hyaluronic acid incorporated into the disks of demineralized bone matrix with 73-90% volume porosity and macropore size from 50 to 650 µm. Dynamics of neoosteogenesis and bone tissue remodeling was investigated histologically at the time points of 12, 21, 48, and 63 days. Diopside particles were evenly spread in the matrix and caused minimal foreign body reaction. In the presence of BMP-2 by the day 63 significant foci of newly formed bone tissue were formed in the implant pores with bone marrow areas, moreover, large areas of demineralized bone matrix in the implant center and maternal bone at the edges were involved in the remodeling. Diopside could be considered as a promising material for introduction into hybrid implants as an effective carrier of BMP-2.

Enterovirus Infection Induces Massive Recruitment of All Isoforms of Small Cellular Arf GTPases to the Replication Organelles.

Enterovirus replication requires the cellular protein GBF1, a guanine nucleotide exchange factor for small Arf GTPases. When activated, Arfs associate with membranes, where they regulate numerous steps of membrane homeostasis. The requirement for GBF1 implies that Arfs are important for replication, but which of the different Arfs function(s) during replication remains poorly understood. Here, we established cell lines expressing each of the human Arfs fused to a fluorescent tag and investigated their behavior during enterovirus infection. Arf1 was the first to be recruited to the replication organelles, where it strongly colocalized with the viral antigen 2B and mature virions but not double-stranded RNA. By the end of the infectious cycle, Arf3, Arf4, Arf5, and Arf6 were also concentrated on the replication organelles. Once on the replication membranes, all Arfs except Arf3 were no longer sensitive to inhibition of GBF1, suggesting that in infected cells they do not actively cycle between GTP- and GDP-bound states. Only the depletion of Arf1, but not other class 1 and 2 Arfs, significantly increased the sensitivity of replication to GBF1 inhibition. Surprisingly, depletion of Arf6, a class 3 Arf, normally implicated in plasma membrane events, also increased the sensitivity to GBF1 inhibition. Together, our results suggest that GBF1-dependent Arf1 activation directly supports the development and/or functioning of the replication complexes and that Arf6 plays a previously unappreciated role in viral replication. Our data reveal a complex pattern of Arf activation in enterovirus-infected cells that may contribute to the resilience of viral replication in different cellular environments.IMPORTANCE Enteroviruses include many known and emerging pathogens, such as poliovirus, enteroviruses 71 and D68, and others. However, licensed vaccines are available only against poliovirus and enterovirus 71, and specific anti-enterovirus therapeutics are lacking. Enterovirus infection induces the massive remodeling of intracellular membranes and the development of specialized domains harboring viral replication complexes, replication organelles. Here, we investigated the roles of small Arf GTPases during enterovirus infection. Arfs control distinct steps in intracellular membrane traffic, and one of the Arf-activating proteins, GBF1, is a cellular factor required for enterovirus replication. We found that all Arfs expressed in human cells, including Arf6, normally associated with the plasma membrane, are recruited to the replication organelles and that Arf1 appears to be the most important Arf for enterovirus replication. These results document the rewiring of the cellular membrane pathways in infected cells and may provide new ways of controlling enterovirus infections.

Liquid-Phase Synthesis of Highly Reactive Rare-Earth Metal Nanoparticles.

The first liquid-phase synthesis of high-quality, small-sized rare-earth metal nanoparticles (1-3 nm)-ranging from lanthanum as one of the largest (187 pm) to scandium as the smallest (161 pm) rare-earth metal-is shown. Size, oxidation state, and reactivity of the nanoparticles are examined (e.g., electron microscopy, electron spectroscopy, X-ray absorption spectroscopy, selected reactions). Whereas the nanoparticles are highly reactive (e.g. in contact to air and water), they are chemically stable as THF suspensions and powders under inert conditions. The reactivity can be controlled to obtain inorganic and metal-organic compounds at room temperature.

Supported Intermetallic PdZn Nanoparticles as Bifunctional Catalysts for the Direct Synthesis of Dimethyl Ether from CO-Rich Synthesis Gas.

The single-step syngas-to-dimethyl ether (STD) process entails economic and technical advantages over the current industrial two-step process. Pd/ZnO-based catalysts have recently emerged as interesting alternatives to currently used Cu/ZnO/Al2 O3 catalysts, but the nature of the active site(s), the reaction mechanism, and the role of Pd and ZnO in the solid catalyst are not well established. Now, Zn-stabilized Pd colloids with a size of 2 nm served as the key building blocks for the methanol active component in bifunctional Pd/ZnO-γ-Al2 O3 catalysts. The catalysts were characterized by combining high-pressure operando X-ray absorption spectroscopy and DFT calculations. The enhanced stability, longevity, and high dimethyl ether selectivity observed makes Pd/ZnO-γ-Al2 O3 an effective alternative system for the STD process compared to Cu/ZnO/γ-Al2 O3 .

Reversible and Irreversible Structural Changes in Cu/ZnO/ZrO2 Catalysts during Methanol Synthesis.

The structure and chemical state of heterogeneous catalysts are closely related to their operational stability. Knowing these relationships as precisely as possible is thus essential for further catalyst development. This work focuses on the deactivation of a Cu/ZnO/ZrO2-type catalyst for methanol synthesis. Experiments were performed in a parallel setup, with which time-dependent changes in the catalyst material can be observed. Elucidation of potential deactivation pathways is described for catalyst aging at different times on stream (0, 50, 935 h). Data from X-ray absorption spectroscopy, X-ray photoelectron spectroscopy, N2 physisorption, and transmission electron microscopy measurements reveal that sintering of Cu0 domains and restructuring within ZnO domains mainly contribute to deactivation. Subsequent reactivation by reduction (in H2/N2) reverts the observed structural changes only to a limited extent. Moreover, this work highlights the participation of ZrO2 as a promoter and reveals redispersion of zirconia after initial reduction.

Aqueous Phase Reforming over Platinum Catalysts on Doped Carbon Supports: Exploring Platinum-Heteroatom Interactions.

A series of platinum catalysts supported on carbon nanofibers with various heteroatom dopings were synthesized to investigate the effect of the local platinum environment on the catalytic activity and selectivity in aqueous phase reforming (APR) of ethylene glycol (EG). Typical carbon dopants such as oxygen, nitrogen, sulfur, phosphorus, and boron were chosen based on their ability to bring acidic or basic functional groups to the carbon surface. In situ X-ray absorption spectroscopy (XAS) was used to identify the platinum oxidation state and platinum species formed during APR of EG through multivariate curve resolution alternating least-squares analysis, observing differences in activity, selectivity, and platinum local environment among the catalysts. The platinum-based catalyst on the nitrogen-doped carbon support demonstrated the most favorable properties for H2 production due to high Pt dispersion and basicity (H2 site time yield 22.7 h-1). Direct Pt-N-O coordination was identified by XAS in this catalyst. The sulfur-doped catalyst presented Pt-S contributions with the lowest EG conversion rate and minimal production of the gas phase components. Boron and phosphorus-doped catalysts showed moderate activity, which was affected by low platinum dispersion on the carbon support. The phosphorus-doped catalyst showed preferential selectivity to alcohols in the liquid phase, associated with the presence of acid sites and Pt-P contributions observed under APR conditions.

Structural Changes of Ni and Ni-Pt Methane Steam Reforming Catalysts During Activation, Reaction, and Deactivation Under Dynamic Reaction Conditions.

Ni-based catalysts are the most widely used materials to produce H2 in large-scale methane steam reformers under stationary conditions. For domestic applications such as fuel cells, H2 production involves the exposure of the catalysts to more dynamic conditions due to the daily startup and shutdown operation mode, making Ni-based catalysts susceptible to oxidation and deactivation. In this context, we report a systematic investigation of the structural changes occurring for monometallic Ni/MgAlOx and bimetallic NiPt/MgAlOx catalysts during methane steam reforming under transient conditions, comprising catalyst activation, operation, and deactivation processes. Besides extensive catalytic tests, the samples prepared by incipient wetness impregnation were characterized by complementary methods, including N2-physisorption, X-ray diffraction, H2-temperature-programmed reduction, and electron microscopy. Next, the structure of the Ni and Pt species was monitored under reaction conditions using time and spatially resolved in situ/operando X-ray absorption spectroscopy. The results obtained show that before catalyst activation by H2-reduction, nickel diffuses into the support lattice and forms mixed oxides with magnesium. In the activated catalysts, Ni is present in the metallic state or alloyed with Pt. A clear beneficial effect of the noble metal addition was identified on both the activity and stability of the bimetallic NiPt/MgAlOx catalyst. In contrast, the pronounced oxidation and reincorporation of Ni into the support lattice were observed for the monometallic sample, and these catalyst deactivation effects are hindered in the bimetallic Ni-Pt catalyst. Overall, the outcome of our study not only helps in understanding the catalyst activation/deactivation processes at an atomic level but also provides the basis for the rational development of improved methane steam reforming catalysts.

Methyl radical chemistry in non-oxidative methane activation over metal single sites.

Molybdenum supported on zeolites has been extensively studied as a catalyst for methane dehydroaromatization. Despite significant progress, the actual intermediates and particularly the first C-C bond formation have not yet been elucidated. Herein we report evolution of methyl radicals during non-oxidative methane activation over molybdenum single sites, which leads selectively to value-added chemicals. Operando X-ray absorption spectroscopy and online synchrotron vacuum ultraviolet photoionization mass spectroscopy in combination with electron microscopy and density functional theory calculations reveal the essential role of molybdenum single sites in the generation of methyl radicals and that the formation rate of methyl radicals is linearly correlated with the number of molybdenum single sites. Methyl radicals transform to ethane in the gas phase, which readily dehydrogenates to ethylene in the absence of zeolites. This is essentially similar to the reaction pathway over the previously reported SiO2 lattice-confined single site iron catalyst. However, the availability of a zeolite, either in a physical mixture or as a support, directs the subsequent reaction pathway towards aromatization within the zeolite confined pores, resulting in benzene as the dominant hydrocarbon product. The findings reveal that methyl radical chemistry could be a general feature for metal single site catalysis regardless of the support (either zeolites MCM-22 and ZSM-5 or SiO2) whereas the reaction over aggregated molybdenum carbide nanoparticles likely facilitates carbon deposition through surface C-C coupling. These findings allow furthering the fundamental insights into non-oxidative methane conversion to value-added chemicals.

Mechanical, Structural, and Biological Characteristics of Polylactide/Wollastonite 3D Printed Scaffolds.

The present work aimed to study the synergistic response of bioresorbable polylactide/bioactive wollastonite scaffolds towards mechanical stability, mesenchymal stromal cell colonization, and antibacterial activity in the physiological environment. Wollastonite was synthesized at 800 °C within 2 h by sol-gel combustion method. The surface area was found to be 1.51 m2/g, and Transmission Electron Microscopy (TEM) micrographs indicated the presence of porous structures. Fused deposition modeling was used to prepare 3D-printed polylactide/wollastonite and polylactide/hydroxyapatite scaffolds. Scanning Electron Microscopy (SEM) micrographs confirmed the interconnected porous structure and complex geometry of the scaffolds. The addition of wollastonite decreased the contact angle of the scaffolds. The mechanical testing of scaffolds examined by computational simulation, as well as machine testing, revealed their non-load-bearing capacity. The chemical constituent of the scaffolds was found to influence the attachment response of different cells on their surface. The incorporation of wollastonite effectively reduced live bacterial attachment, whereas the colonization of mesenchymal cells was improved. This observation confirms polylactide/wollastonite scaffold possesses both bactericidal as well as cytocompatible properties. Thus, the risk of peri-implant bacterial film formation can be prevented, and the biological fixation of the scaffold at the defect site can be enhanced by utilizing these composites.

Effect of recombinant BMP-2 and erythropoietin on osteogenic properties of biomimetic PLA/PCL/HA and PHB/HA scaffolds in critical-size cranial defects model.

Osteoplastic materials PLA/PCL/HA and PHB/HA and scaffolds with a highly porous structure based on them with potential applications in regenerative medicine have been obtained by solvent casting with thermopressing and salt leaching for PLA-based samples and solid-state mixing with subsequent thermopressing and salt leaching for PHB-based samples. The scaffolds were characterized by SEM-EDX, DSC, FTIR spectroscopy, mechanical tests in compression, measurement of the contact angle, in vitro studies, including loading by recombinant BMP-2 and EPO and their release kinetics, and in vivo studies on a model of regeneration of critical-sized cranial defects in mice. Biomimetic scaffolds with micropores sizes ranged from 300 to 500 µm and volume porosity of 70% imitate trabecular bone's structure and have increased hydrophilicity to achieve osteoconductive properties. Mechanical characteristics correspond to native trabecular bone. Elastic modulus - key mechanical characteristics of bone implants - showed the values of 0.15 ± 0.04 and 0.18 ± 0.08 GPa for PLA/PCL/HA and PHB/HA scaffolds, respectively. Both materials have high biocompatibility and can be used together with recombinant proteins BMP-2 and EPO. Introduction of BMP-2 leads to induction of new bone formation, introduction of EPO results in increased angiogenesis in the implantation area. The obtained scaffolds with recombinant proteins can be used as bone implants for reconstruction of defects of lightly or non-loaded bones.

Genetic Variations and Haplotypic Diversity in the Myostatin Gene of Different Cattle Breeds in Russia.

The myostatin gene (MSTN) in cattle has a number of polymorphisms associated with increased muscle mass. The aim of the current study was to determine the haplotype frequencies of F94L and nt821(del11) MSTN polymorphisms among cattle bred for meat in Russia, using DNA analysis. Using the earlier created test systems based on the AS-PCR and PCR-RFLP methods, six populations of Aberdeen Angus (n = 684), two populations of Limousin (n = 54), one population of Simmental (n = 55), and one population of Belgian Blue (n = 137) belonging to Russian farms were genotyped on nt821(del11) and F94LMSTN polymorphisms. The animal carriers of the mutant allele of nt821(del11)MSTN associated with the double-muscling genetic defect were found in one Aberdeen Angus population at a frequency of 2.18%, but were not found in the Limousin and Simmental populations. However, 100% of the Belgian Blue population were heterozygous carriers of nt821(del11)MSTN. The frequencies of the A allele F94LMSTN desirable for productivity traits in the Limousin populations were the highest and accounted for 0.97 and 1 in populations one and two, while in the Aberdeen Angus, Simmental, and Belgian Blue populations, these figures were considerably lower at 0.04-0.08, depending on the population. The obtained data show the high genetic potential of Russian beef cattle, and facilitate an improvement in meat productivity by preserving the health of animals.

Genomic Regions and Candidate Genes Linked to Capped Hock in Pig.

Capped hock affects the exterior of pedigree pigs, making them unsalable and resulting in a negative impact on the efficiency of pig-breeding centers. The purpose of this paper was to carry out pilot studies aimed at finding genomic regions and genes linked to the capped hock in pigs. The studies were carried out on Landrace pigs (n = 75) and Duroc pigs (n = 70). To identify genomic regions linked to capped hock in pigs, we used smoothing FST statistics. Genotyping was performed with GeneSeek® GGP Porcine HD Genomic Profiler v1 (Illumina Inc, USA). The research results showed 70 SNPs linked to capped hock in Landrace (38 SNPs) and Duroc (32 SNPs). The identified regions overlapped with QTLs related with health traits (blood parameters) and meat and carcass traits (fatness). In total, 31 genes were identified (i.e., 17 genes in Landrace, 14 genes in Durocs). Three genes appeared in both the Landrace and Duroc groups, including A2ML1 (SSC5), ROBO2 (SSC13), and MSI1 (SSC14). We identified genomic regions directly or indirectly linked to capped hock, which thus might contribute to identifying genetic variants and using them as genetic markers in pig breeding.

Interaction of Poliovirus Capsid Proteins with the Cellular Autophagy Pathway.

The capsid precursor P1 constitutes the N-terminal part of the enterovirus polyprotein. It is processed into VP0, VP3, and VP1 by the viral proteases, and VP0 is cleaved autocatalytically into VP4 and VP2. We observed that poliovirus VP0 is recognized by an antibody against a cellular autophagy protein, LC3A. The LC3A-like epitope overlapped the VP4/VP2 cleavage site. Individually expressed VP0-EGFP and P1 strongly colocalized with a marker of selective autophagy, p62/SQSTM1. To assess the role of capsid proteins in autophagy development we infected different cells with poliovirus or encapsidated polio replicon coding for only the replication proteins. We analyzed the processing of LC3B and p62/SQSTM1, markers of the initiation and completion of the autophagy pathway and investigated the association of the viral antigens with these autophagy proteins in infected cells. We observed cell-type-specific development of autophagy upon infection and found that only the virion signal strongly colocalized with p62/SQSTM1 early in infection. Collectively, our data suggest that activation of autophagy is not required for replication, and that capsid proteins contain determinants targeting them to p62/SQSTM1-dependent sequestration. Such a strategy may control the level of capsid proteins so that viral RNAs are not removed from the replication/translation pool prematurely.