The sequential steps in the alphavirus membrane fusion pathway have been postulated based on the prefusion and postfusion crystal structures of the viral fusion protein E1 in conjunction with biochemical studies. However, the molecular structures of the hypothesized fusion intermediates have remained obscure due to difficulties inherent in the dynamic nature of the process. We developed an experimental system that uses liposomes as the target membrane to capture Sindbis virus, a prototypical alphavirus, in its membrane-binding form at pH 6.4. Cryoelectron micrograph analyses and 3D reconstructions showed that the virus retains its overall icosahedral structure at this mildly acidic pH, except in the membrane-binding region, where monomeric E1 associates with the target membrane and the E2 glycoprotein retains its original trimeric organization. The remaining E2 trimers may hinder E1 homotrimerization and are a potential target for antiviral drugs.
The double ionization of helium by electron impact for $106\phantom{\rule{0.3em}{0ex}}\mathrm{eV}$ incident energy was studied in a kinematically complete experiment by using a reaction microscope. The pattern of the angular correlation of the three emitted electrons was analyzed by selecting different values of the recoil ion longitudinal momentum. The Wannier predicted geometry appears when the recoil ion carries the full initial projectile momentum. It was found that at this low impact energy, the outgoing electrons still remember the initial-state collision information.
ABSTRACT The emergence of novel variants of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) continues to pose an ongoing challenge for global public health services, highlighting the urgent need for effective therapeutic interventions. Neutralizing monoclonal antibodies (mAbs) are a major therapeutic strategy for the treatment of COVID-19 and other viral diseases. In this study, we employed hybridoma technology to generate mAbs that target the BA.5 receptor-binding domain (RBD) of the SARS-CoV-2 spike protein. Through a comprehensive screening process, we identified four mAbs capable of effectively neutralizing BA.5, XBB.1.16, and related variant infections in vitro , among which ORB10 was found to neutralize BA.5 variants with a plaque reduction neutralization test (PRNT 50 ) of 8.7 ng/mL. Additionally, competitive binding assays, sequencing of heavy and light chain variable regions, and binding kinetics characterization provided insights into the epitopes and binding affinities of the identified mAbs. Moreover, in vivo experiments in the K18-hACE2 mouse model demonstrated the protective efficacy of ORB10 against both BA.5 and XBB.1.16 variants. Finally, cryo-electron microscopy structural analysis of the ORB10–RBD complex identified key residues involved in the antibody–antigen interactions, providing insights into the molecular mechanisms of neutralization and immune escape of SARS-CoV-2 Omicron variants from mAbs. IMPORTANCE The ongoing evolution of SARS-CoV-2 has led to the emergence of variants capable of evading immune responses elicited by natural infection and vaccination, especially the highly transmissible and immune-evasive Omicron variants. This study generated and characterized a panel of monoclonal antibodies (mAbs) specifically targeting the RBD of the Omicron BA.5 variant, of which the ORB10 showed efficacy against Omicron BA.5 and XBB.1.16 variants both in vitro and in vivo . Cryo-EM structural analysis further elucidated the binding epitope interactions and neutralization mechanism between ORB10 and the BA.5 RBD protein. This study enhances our understanding of antibody-mediated neutralization of SARS-CoV-2 and provides valuable insights into the development of effective therapeutic strategies to combat ongoing SARS-CoV-2 variant infections.
Abstract Banna virus (BAV) is the prototype Seadornavirus , a class of reoviruses for which there has been little structural study. Here, we report atomic cryo-EM structures of three states of BAV virions—surrounded by 120 spikes (full virions), 60 spikes (partial virions), or no spikes (cores). BAV cores are double-layered particles similar to the cores of other non-turreted reoviruses, except for an additional protein component in the outer capsid shell, VP10. VP10 was identified to be a cementing protein that plays a pivotal role in the assembly of BAV virions by directly interacting with VP2 (inner capsid), VP8 (outer capsid), and VP4 (spike). Viral spikes (VP4/VP9 heterohexamers) are situated on top of VP10 molecules in full or partial virions. Asymmetrical electrostatic interactions between VP10 monomers and VP4 trimers are disrupted by high pH treatment, which is thus a simple way to produce BAV cores. Low pH treatment of BAV virions removes only the flexible receptor binding protein VP9 and triggers significant conformational changes in the membrane penetration protein VP4. BAV virions adopt distinct spatial organization of their surface proteins compared with other well-studied reoviruses, suggesting that BAV may have a unique mechanism of penetration of cellular endomembranes.
Abstract HSP gp96‐based vaccines have been trialled in rodent models and, more recently, in humans. Better understanding of gp96's immunomodulatory role will help with the design of more effective strategies for treatment of cancer and infectious diseases. In this study, we monitored the activities of T cells and activation of Treg in BABL/c mice after immunization using different doses of gp96 as adjuvant. We found that co‐injection of gp96 simultaneously stimulated both CTL and Treg activity. Activation of CTL at low dose was far more pronounced than Treg activation. Treg population and suppression increased with gp96 dose, eventually abrogating the T‐cell response induced by immunization. Low‐dose cyclophosphamide treatment could restore the T‐cell responses lost after high‐dose gp96 adjuvant injection by suppression of Treg activation. We further examined the effect of different doses of gp96 or N355 peptide administration on tumor rejection. Our results provide new insights into the mechanisms of gp96‐mediated balance between regulatory and responder T cells, which may facilitate future development of an effective gp96‐based therapeutic vaccine.
Presently, integrating multi-omics information into a prediction model has become a ameliorate strategy for genomic selection to improve genomic prediction accuracy. Here, we set the genomic and transcriptomic data as the training population data, using BSLMM, TWAS, and eQTL mapping to prescreen features according to |β∘b|>0, top1% percent of phenotypic variation explained (PVE), expression-associated single nucleotide polymorphisms (eSNPs), and egenes (false discovery rate (FDR) < 0.01), where these loci were set as extra fixed effects (named GBLUP-Fix) and random effects (GFBLUP) to improve the prediction accuracy in the validation population, respectively. The results suggested that both GBLUP-Fix and GFBLUP models could improve the accuracy of longissimus dorsi muscle (LDM), water holding capacity WHC, shear force SF, and pH in Huaxi cattle on average from 2.14% to 8.69%, especially the improvement of GFBLUP-TWAS over GBLUP was 13.66% for SF. These methods also captured more genetic variance than GBLUP. Our study confirmed that multi-omics-assisted large-effects loci prescreening could improve the accuracy of genomic prediction.
Abstract Incorporating the genotypic and phenotypic of the correlated traits into the multi-trait model can significantly improve the prediction accuracy of the target trait in animal and plant breeding, as well as human genetics. However, in most cases, the phenotypic information of the correlated and target trait of the individual to be evaluated was null simultaneously, particularly for the newborn. Therefore, we propose a machine learning framework, MAK, to improve the prediction accuracy of the target trait by constructing the multi-target ensemble regression chains and selecting the assistant trait automatically, which predicted the genomic estimated breeding values of the target trait using genotypic information only. The prediction ability of MAK was significantly more robust than the genomic best linear unbiased prediction, BayesB, BayesRR and the multi trait Bayesian method in the four real animal and plant datasets, and the computational efficiency of MAK was roughly 100 times faster than BayesB and BayesRR.
Crimean-Congo hemorrhagic fever virus (CCHFV) is a highly pathogenic tick-borne virus, prevalent in more than 30 countries worldwide. Human infection by this virus leads to severe illness, with an average case fatality of 40%. There is currently no approved vaccine or drug to treat the disease. Neutralizing antibodies are a promising approach to treat virus infectious diseases. This study generated 37 mouse-derived specific monoclonal antibodies against CCHFV Gc subunit. Neutralization assays using pseudotyped virus and authentic CCHFV identified Gc8, Gc13, and Gc35 as neutralizing antibodies. Among them, Gc13 had the highest neutralizing activity and binding affinity with CCHFV Gc. Consistently, Gc13, but not Gc8 or Gc35, showed in vivo protective efficacy (62.5% survival rate) against CCHFV infection in a lethal mouse infection model. Further characterization studies suggested that Gc8 and Gc13 may recognize a similar, linear epitope in domain II of CCHFV Gc, while Gc35 may recognize a different epitope in Gc. Cryo-electron microscopy of Gc-Fab complexes indicated that both Gc8 and Gc13 bind to the conserved fusion loop region and Gc13 had stronger interactions with sGc-trimers. This was supported by the ability of Gc13 to block CCHFV GP-mediated membrane fusion. Overall, this study provides new therapeutic strategies to treat CCHF and new insights into the interaction between antibodies with CCHFV Gc proteins.
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