Immunization-Elicited Broadly Protective Antibody Reveals Ebolavirus Fusion Loop as a Site of Vulnerability.

While neutralizing antibodies are highly effective against ebolavirus infections, current experimental ebolavirus vaccines primarily elicit species-specific antibody responses. Here, we describe an immunization-elicited macaque antibody (CA45) that clamps the internal fusion loop with the N terminus of the ebolavirus glycoproteins (GPs) and potently neutralizes Ebola, Sudan, Bundibugyo, and Reston viruses. CA45, alone or in combination with an antibody that blocks receptor binding, provided full protection against all pathogenic ebolaviruses in mice, guinea pigs, and ferrets. Analysis of memory B cells from the immunized macaque suggests that elicitation of broadly neutralizing antibodies (bNAbs) for ebolaviruses is possible but difficult, potentially due to the rarity of bNAb clones and their precursors. Unexpectedly, germline-reverted CA45, while exhibiting negligible binding to full-length GP, bound a proteolytically remodeled GP with picomolar affinity, suggesting that engineered ebolavirus vaccines could trigger rare bNAb precursors more robustly. These findings have important implications for developing pan-ebolavirus vaccine and immunotherapeutic cocktails.

IgG3 regulates tissue-like memory B cells in HIV-infected individuals.

Immunoglobulin G3 (IgG3) has an uncertain role in the response to infection with and vaccination against human immunodeficiency virus (HIV). Here we describe a regulatory role for IgG3 in dampening the immune system-activating effects of chronic HIV viremia on B cells. Secreted IgG3 was bound to IgM-expressing B cells in vivo in HIV-infected chronically viremic individuals but not in early-viremic or aviremic individuals. Tissue-like memory (TLM) B cells, a population expanded by persistent HIV viremia, bound large amounts of IgG3. IgG3 induced clustering of B cell antigen receptors (BCRs) on the IgM+ B cells, which was mediated by direct interactions between soluble IgG3 and membrane IgM of the BCR (IgM-BCR). The inhibitory IgG receptor CD32b (FcγRIIb), complement component C1q and inflammatory biomarker CRP contributed to the binding of secreted IgG3 onto IgM-expressing B cells of HIV-infected individuals. Notably, IgG3-bound TLM B cells were refractory to IgM-BCR stimulation, thus demonstrating that IgG3 can regulate B cells during chronic activation of the immune system.

Spliceosome component PHD finger 5A is essential for early B lymphopoiesis.

The spliceosome, a multi-megadalton ribonucleoprotein complex, is essential for pre-mRNA splicing in the nucleus and ensuring genomic stability. Its precise and dynamic assembly is pivotal for its function. Spliceosome malfunctions can lead to developmental abnormalities and potentially contribute to tumorigenesis. The specific role of the spliceosome in B cell development is poorly understood. Here, we reveal that the spliceosomal U2 snRNP component PHD finger protein 5A (Phf5a) is vital for early B cell development. Loss of Phf5a results in pronounced defects in B cell development, causing an arrest at the transition from pre-pro-B to early pro-B cell stage in the bone marrow of mutant mice. Phf5a-deficient B cells exhibit impaired immunoglobulin heavy (IgH) chain expression due to defective V-to-DJ gene rearrangement. Mechanistically, our findings suggest that Phf5a facilitates IgH gene rearrangement by regulating the activity of recombination-activating gene endonuclease and influencing chromatin interactions at the Igh locus.

Identification, structural, and biophysical characterization of a positive modulator of human Kv3.1 channels.

Voltage-gated potassium channels (Kv) are tetrameric membrane proteins that provide a highly selective pathway for potassium ions (K+) to diffuse across a hydrophobic cell membrane. These unique voltage-gated cation channels detect changes in membrane potential and, upon activation, help to return the depolarized cell to a resting state during the repolarization stage of each action potential. The Kv3 family of potassium channels is characterized by a high activation potential and rapid kinetics, which play a crucial role for the fast-spiking neuronal phenotype. Mutations in the Kv3.1 channel have been shown to have implications in various neurological diseases like epilepsy and Alzheimer's disease. Moreover, disruptions in neuronal circuitry involving Kv3.1 have been correlated with negative symptoms of schizophrenia. Here, we report the discovery of a novel positive modulator of Kv3.1, investigate its biophysical properties, and determine the cryo-EM structure of the compound in complex with Kv3.1. Structural analysis reveals the molecular determinants of positive modulation in Kv3.1 channels by this class of compounds and provides additional opportunities for rational drug design for the treatment of associated neurological disorders.

Key gp120 Glycans Pose Roadblocks to the Rapid Development of VRC01-Class Antibodies in an HIV-1-Infected Chinese Donor.

VRC01-class antibodies neutralize diverse HIV-1 strains by targeting the conserved CD4-binding site. Despite extensive investigations, crucial events in the early stage of VRC01 development remain elusive. We demonstrated how VRC01-class antibodies emerged in a Chinese donor by antigen-specific single B cell sorting, structural and functional studies, and longitudinal antibody and virus repertoire analyses. A monoclonal antibody DRVIA7 with modest neutralizing breadth was isolated that displayed a subset of VRC01 signatures. X-ray and EM structures revealed a VRC01-like angle of approach, but less favorable interactions between the DRVIA7 light-chain CDR1 and the N terminus with N276 and V5 glycans of gp120. Although the DRVIA7 lineage was unable to acquire broad neutralization, longitudinal analysis revealed a repertoire-encoded VRC01 light-chain CDR3 signature and VRC01-like neutralizing heavy-chain precursors that rapidly matured within 2 years. Thus, light chain accommodation of the glycan shield should be taken into account in vaccine design targeting this conserved site of vulnerability.

Early human B cell signatures of the primary antibody response to mRNA vaccination.

Messenger RNA (mRNA) vaccines against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are highly effective at inducing protective immunity. However, weak antibody responses are seen in some individuals, and cellular correlates of immunity remain poorly defined, especially for B cells. Here we used unbiased approaches to longitudinally dissect primary antibody, plasmablast, and memory B cell (MBC) responses to the two-dose mRNA-1273 vaccine in SARS-CoV-2-naive adults. Coordinated immunoglobulin A (IgA) and IgG antibody responses were preceded by bursts of spike-specific plasmablasts after both doses but earlier and more intensely after dose 2. While antibody and B cell cellular responses were generally robust, they also varied within the cohort and decreased over time after a dose-2 peak. Both antigen-nonspecific postvaccination plasmablast frequency after dose 1 and their spike-specific counterparts early after dose 2 correlated with subsequent antibody levels. This correlation between early plasmablasts and antibodies remained for titers measured at 6 months after vaccination. Several distinct antigen-specific MBC populations emerged postvaccination with varying kinetics, including two MBC populations that correlated with 2- and 6-month antibody titers. Both were IgG-expressing MBCs: one less mature, appearing as a correlate after the first dose, while the other MBC correlate showed a more mature and resting phenotype, emerging as a correlate later after dose 2. This latter MBC was also a major contributor to the sustained spike-specific MBC response observed at month 6. Thus, these plasmablasts and MBCs that emerged after both the first and second doses with distinct kinetics are potential determinants of the magnitude and durability of antibodies in response to mRNA-based vaccination.

Glucose uptake from the rhizosphere mediated by MdDOF3-MdHT1.2 regulates drought resistance in apple.

In plants under drought stress, sugar content in roots increases, which is important for drought resistance. However, the molecular mechanisms for controlling the sugar content in roots during response to drought remain elusive. Here, we found that the MdDOF3-MdHT1.2 module-mediated glucose influx into the root is essential for drought resistance in apple (Malus × domestica). Drought induced glucose uptake from the rhizosphere and up-regulated the transcription of hexose transporter MdHT1.2. Compared with the wild-type plants, overexpression of MdHT1.2 promoted glucose uptake from the rhizosphere, thereby facilitating sugar accumulation in root and enhancing drought resistance, whereas silenced plants showed the opposite phenotype. Furthermore, ATAC-seq, RNA-seq and biochemical analysis demonstrated that MdDOF3 directly bound to the promoter of MdHT1.2 and was strongly up-regulated under drought. Overexpression of MdDOF3 in roots improved MdHT1.2-mediated glucose transport capacity and enhanced plant resistance to drought, but MdDOF3-RNAihr apple plants showed the opposite phenotype. Moreover, overexpression of MdDOF3 in roots did not attenuate drought sensitivity in MdHT1.2-RNAi plants, which was correlated with a lower glucose uptake capacity and glucose content in root. Collectively, our findings deciphered the molecular mechanism through which glucose uptake from the rhizosphere is mediated by MdDOF3-MdHT1.2, which acts to modulate sugar content in root and promote drought resistance.

Uptake of glucose from the rhizosphere, mediated by apple MdHT1.2, regulates carbohydrate allocation.

Plant roots can absorb sugars from the rhizosphere, which reduces the consumption of carbon derived from photosynthesis. However, the underlying mechanisms that roots use to control sugar absorption from soil are poorly understood. Here, we identified an apple (Malus × domestica Borkh.) hexose transporter, MdHT1.2, that functions on the root epidermis to absorb glucose (Glc) from the rhizosphere. Based on RNA-seq data, MdHT1.2 showed the highest expression level among 29 MdHT genes in apple roots. Biochemical analyses demonstrated that MdHT1.2 was mainly expressed in the epidermal cells of fine roots, and its protein was located on the plasma membrane. The roots of transgenic apple and Solanum lycopersicum lines overexpressing MdHT1.2 had an increased capability to absorb Glc when fed with [13C]-labeled Glc or 2-NBDG, whereas silencing MdHT1.2 in apple showed the opposite results. Further studies established that MdHT1.2-mediated Glc absorption from the rhizosphere changed the carbon assimilate allocation between apple shoot and root, which regulated plant growth. Additionally, a grafting experiment in tomato confirmed that increasing the Glc uptake capacity in the root overexpressing MdHT1.2 could facilitate carbohydrate partitioning to the fruit. Collectively, our study demonstrated that MdHT1.2 functions on the root epidermis to absorb rhizospheric Glc, which regulates the carbohydrate allocation for plant growth and fruit sugar accumulation.

Mettl14-Mediated m6A Modification Is Essential for Germinal Center B Cell Response.

The germinal center (GC) response is essential for generating memory B and long-lived Ab-secreting plasma cells during the T cell-dependent immune response. In the GC, signals via the BCR and CD40 collaboratively promote the proliferation and positive selection of GC B cells expressing BCRs with high affinities for specific Ags. Although a complex gene transcriptional regulatory network is known to control the GC response, it remains elusive how the positive selection of GC B cells is modulated posttranscriptionally. In this study, we show that methyltransferase like 14 (Mettl14)-mediated methylation of adenosines at the position N 6 of mRNA (N 6-methyladenosine [m6A]) is essential for the GC B cell response in mice. Ablation of Mettl14 in B cells leads to compromised GC B cell proliferation and a defective Ab response. Interestingly, we unravel that Mettl14-mediated m6A regulates the expression of genes critical for positive selection and cell cycle regulation of GC B cells in a Ythdf2-dependent but Myc-independent manner. Furthermore, our study reveals that Mettl14-mediated m6A modification promotes mRNA decay of negative immune regulators, such as Lax1 and Tipe2, to upregulate genes requisite for GC B cell positive selection and proliferation. Thus, our findings suggest that Mettl14-mediated m6A modification plays an essential role in the GC B cell response.

RNA-Editing Enzyme ADAR1 p150 Isoform Is Critical for Germinal Center B Cell Response.

Adenosine deaminase acting on RNA (ADAR)1 is the principal enzyme for adenosine-to-inosine editing, an RNA modification-avoiding cytosolic nucleic acid sensor's activation triggered by endogenous dsRNAs. Two ADAR1 isoforms exist in mammals, a longer IFN-inducible and mainly cytoplasm-localized p150 isoform and a shorter constitutively expressed and primarily nucleus-localized p110 isoform. Studies of ADAR1 mutant mice have demonstrated that ADAR1 is essential for multiple physiological processes, including embryonic development, innate immune response, and B and T lymphocyte development. However, it remained unknown whether ADAR1 plays a role in the humoral immune response. In this study, we conditionally delete Adar1 in activated B cells and show that ADAR1-deficient mice have a defective T cell-dependent Ab response and diminished germinal center (GC) B cells. Using various double mutant mice concurrently deficient in ADAR1 and different downstream dsRNA sensors, we demonstrate that ADAR1 regulates the GC response by preventing hyperactivation of the melanoma differentiation-associated protein 5 (MDA5) but not the protein kinase R or RNase L pathway. We also show that p150 is exclusively responsible for ADAR1's function in the GC response, and the p110 isoform cannot substitute for the p150's role, even when p110 is constitutively expressed in the cytoplasm. We further demonstrated that the dsRNA-binding but not the RNA-editing activity is required for ADAR1's function in the GC response. Thus, our data suggest that the ADAR1 p150 isoform plays a crucial role in regulating the GC B cell response.

Research on Sea State Signal Recognition Based on Beluga Whale Optimization-Slope Entropy and One Dimensional-Convolutional Neural Network.

This study introduces a novel nonlinear dynamic analysis method, known as beluga whale optimization-slope entropy (BWO-SlEn), to address the challenge of recognizing sea state signals (SSSs) in complex marine environments. A method of underwater acoustic signal recognition based on BWO-SlEn and one-dimensional convolutional neural network (1D-CNN) is proposed. Firstly, particle swarm optimization-slope entropy (PSO-SlEn), BWO-SlEn, and Harris hawk optimization-slope entropy (HHO-SlEn) were used for feature extraction of noise signal and SSS. After 1D-CNN classification, BWO-SlEn were found to have the best recognition effect. Secondly, fuzzy entropy (FE), sample entropy (SE), permutation entropy (PE), and dispersion entropy (DE) were used to extract the signal features. After 1D-CNN classification, BWO-SlEn and 1D-CNN were found to have the highest recognition rate compared with them. Finally, compared with the other six recognition methods, the recognition rates of BWO-SlEn and 1D-CNN for the noise signal and SSS are at least 6% and 4.75% higher, respectively. Therefore, the BWO-SlEn and 1D-CNN recognition methods proposed in this paper are more effective in the application of SSS recognition.

Detection and Feature Extraction in Acoustic Sensor Signals.

Our advances in detection and feature extraction in the processing of acoustic signals allow us to capture more information about a target and extract features with separability [...].

A Dual-Optimization Fault Diagnosis Method for Rolling Bearings Based on Hierarchical Slope Entropy and SVM Synergized with Shark Optimization Algorithm.

Slope entropy (SlopEn) has been widely applied in fault diagnosis and has exhibited excellent performance, while SlopEn suffers from the problem of threshold selection. Aiming to further enhance the identifying capability of SlopEn in fault diagnosis, on the basis of SlopEn, the concept of hierarchy is introduced, and a new complexity feature, namely hierarchical slope entropy (HSlopEn), is proposed. Meanwhile, to address the problems of the threshold selection of HSlopEn and a support vector machine (SVM), the white shark optimizer (WSO) is applied to optimize both HSlopEn and an SVM, and WSO-HSlopEn and WSO-SVM are proposed, respectively. Then, a dual-optimization fault diagnosis method for rolling bearings based on WSO-HSlopEn and WSO-SVM is put forward. We conducted measured experiments on single- and multi-feature scenarios, and the experimental results demonstrated that whether single-feature or multi-feature, the WSO-HSlopEn and WSO-SVM fault diagnosis method has the highest recognition rate compared to other hierarchical entropies; moreover, under multi-features, the recognition rates are all higher than 97.5%, and the more features we select, the better the recognition effect. When five nodes are selected, the highest recognition rate reaches 100%.

Variable-Step Multiscale Fuzzy Dispersion Entropy: A Novel Metric for Signal Analysis.

Fuzzy dispersion entropy (FuzDE) is a newly proposed entropy metric, which combines the superior characteristics of fuzzy entropy (FE) and dispersion entropy (DE) in signal analysis. However, FuzDE only reflects the feature from the original signal, which ignores the hidden information on the time scale. To address this problem, we introduce variable-step multiscale processing in FuzDE and propose variable-step multiscale FuzDE (VSMFuzDE), which realizes the characterization of abundant scale information, and is not limited by the signal length like the traditional multiscale processing. The experimental results for both simulated signals show that VSMFuzDE is more robust, more sensitive to dynamic changes in the chirp signal, and has more separability for noise signals; in addition, the proposed VSMFuzDE displays the best classification performance in both real-world signal experiments compared to the other four entropy metrics, the highest recognition rates of the five gear signals and four ship-radiated noises reached 99.2% and 100%, respectively, which achieves the accurate identification of two different categories of signals.

Volumetric segmentation of white matter tracts with label embedding.

Convolutional neural networks have achieved state-of-the-art performance for white matter (WM) tract segmentation based on diffusion magnetic resonance imaging (dMRI). However, the segmentation can still be difficult for challenging WM tracts with thin bodies or complicated shapes; the segmentation is even more problematic in challenging scenarios with reduced data quality or domain shift between training and test data, which can be easily encountered in clinical settings. In this work, we seek to improve the segmentation of WM tracts, especially for challenging WM tracts in challenging scenarios. In particular, our method is based on volumetric WM tract segmentation, where voxels are directly labeled without performing tractography. To improve the segmentation, we exploit the characteristics of WM tracts that different tracts can cross or overlap and revise the network design accordingly. Specifically, because multiple tracts can co-exist in a voxel, we hypothesize that the different tract labels can be correlated. The tract labels at a single voxel are concatenated as a label vector, the length of which is the number of tract labels. Due to the tract correlation, this label vector can be projected into a lower-dimensional space-referred to as the embedded space-for each voxel, which allows the segmentation network to solve a simpler problem. By predicting the coordinate in the embedded space for the tracts at each voxel and subsequently mapping the coordinate to the label vector with a reconstruction module, the segmentation result can be achieved. To facilitate the learning of the embedded space, an auxiliary label reconstruction loss is integrated with the segmentation accuracy loss during network training, and network training and inference are end-to-end. Our method was validated on two dMRI datasets under various settings. The results show that the proposed method improves the accuracy of WM tract segmentation, and the improvement is more prominent for challenging tracts in challenging scenarios.

Prominent Neutralizing Antibody Response Targeting the Ebolavirus Glycoprotein Subunit Interface Elicited by Immunization.

The severe death toll caused by the recent outbreak of Ebola virus disease reinforces the importance of developing ebolavirus prevention and treatment strategies. Here, we have explored the immunogenicity of a novel immunization regimen priming with vesicular stomatitis virus particles bearing Sudan Ebola virus (SUDV) glycoprotein (GP) that consists of GP1 & GP2 subunits and boosting with soluble SUDV GP in macaques, which developed robust neutralizing antibody (nAb) responses following immunizations. Moreover, EB46, a protective nAb isolated from one of the immune macaques, is found to target the GP1/GP2 interface, with GP-binding mode and neutralization mechanism similar to a number of ebolavirus nAbs from human and mouse, indicating that the ebolavirus GP1/GP2 interface is a common immunological target in different species. Importantly, selected immune macaque polyclonal sera showed nAb specificity similar to EB46 at substantial titers, suggesting that the GP1/GP2 interface region is a viable target for ebolavirus vaccine.Importance: The elicitation of sustained neutralizing antibody (nAb) responses against diverse ebolavirus strains remains as a high priority for the vaccine field. The most clinically advanced rVSV-ZEBOV vaccine could elicit moderate nAb responses against only one ebolavirus strain, EBOV, among the five ebolavirus strains, which last less than 6 months. Boost immunization strategies are desirable to effectively recall the rVSV vector-primed nAb responses to prevent infections in prospective epidemics, while an in-depth understanding of the specificity of immunization-elicited nAb responses is essential for improving vaccine performance. Here, using non-human primate animal model, we demonstrated that booster immunization with a stabilized trimeric soluble form of recombinant glycoprotein derived from the ebolavirus Sudan strain following the priming rVSV vector immunization led to robust nAb responses that substantially map to the subunit interface of ebolavirus glycoprotein, a common B cell repertoire target of multiple species including primates and rodents.

HIV-1 gp120-CD4-Induced Antibody Complex Elicits CD4 Binding Site-Specific Antibody Response in Mice.

Elicitation of broadly neutralizing Ab (bNAb) responses toward the conserved HIV-1 envelope (Env) CD4 binding site (CD4bs) by vaccination is an important goal for vaccine development and yet to be achieved. The outcome of previous immunogenicity studies suggests that the limited accessibility of the CD4bs and the presence of predominant nonneutralizing determinants (nND) on Env may impede the elicitation of bNAbs and their precursors by vaccination. In this study, we designed a panel of novel immunogens that 1) preferentially expose the CD4bs by selective elimination of glycosylation sites flanking the CD4bs, and 2) minimize the nND immune response by engineering fusion proteins consisting of gp120 Core and one or two CD4-induced (CD4i) mAbs for masking nND epitopes, referred to as gp120-CD4i fusion proteins. As expected, the fusion proteins possess improved antigenicity with retained affinity for VRC01-class, CD4bs-directed bNAbs and dampened affinity for nonneutralizing Abs. We immunized C57BL/6 mice with these fusion proteins and found that overall the fusion proteins elicit more focused CD4bs Ab response than prototypical gp120 Core by serological analysis. Consistently, we found that mice immunized with selected gp120-CD4i fusion proteins have higher frequencies of germinal center-activated B cells and CD4bs-directed memory B cells than those inoculated with parental immunogens. We isolated three mAbs from mice immunized with selected gp120-CD4i fusion proteins and found that their footprints on Env are similar to VRC01-class bNAbs. Thus, using gp120-CD4i fusion proteins with selective glycan deletion as immunogens could focus Ab response toward CD4bs epitope.

Entropy and Information Theory in Acoustics.

Acoustics is one of the most studied fields in the 21st century, encompassing underwater acoustics, architectural acoustics, engineering acoustics, physical acoustics, environmental acoustics, psychological acoustics, signal processing in acoustics, and so on [...].

Optimized Ship-Radiated Noise Feature Extraction Approaches Based on CEEMDAN and Slope Entropy.

Slope entropy (Slopen) has been demonstrated to be an excellent approach to extracting ship-radiated noise signals (S-NSs) features by analyzing the complexity of the signals; however, its recognition ability is limited because it extracts the features of undecomposed S-NSs. To solve this problem, in this study, we combined complete ensemble empirical mode decomposition with adaptive noise (CEEMDAN) to explore the differences of Slopen between the intrinsic mode components (IMFs) of the S-NSs and proposed a single-IMF optimized feature extraction approach. Aiming to further enhance its performance, the optimized combination of dual-IMFs was selected, and a dual-IMF optimized feature extraction approach was also proposed. We conducted three experiments to demonstrate the effectiveness of CEEMDAN, Slopen, and the proposed approaches. The experimental and comparative results revealed both of the proposed single- and dual-IMF optimized feature extraction approaches based on Slopen and CEEMDAN to be more effective than the original ship signal-based and IMF-based feature extraction approaches.

Investigation on Hydrate Growth at the Oil-Water Interface: In the Presence of Wax and Surfactant.

Natural gas hydrates can readily form in deep-water-oil production processes and pose a great threat to subsea pipeline flow assurance. The usage of surfactants and hydrate antiagglomerants is a common strategy to prevent hydrate hazards. In water/wax-containing oil systems, hydrate coexisting with wax could lead to more complex and risky transportation conditions. Moreover, the effectiveness of surfactants and hydrate antiagglomerants in the presence of wax should be further evaluated. In this work, for the purpose of investigating how wax and surfactants could affect hydrate growth at the oil-water interface, a series of microexperiments was conducted in an atmospheric visual cell where the nucleation and growth of hydrates took place on a water droplet surrounded by wax-containing oils. On the basis of the experimental phenomena observed using a microscope, the formation of a hydrate shell by lateral growth, the collapse of a water droplet after hydrate initial formation, and the formation of hollow-conical hydrate crystals were identified. These experimental phenomena were closely related to the concentration of wax and surfactant used in each case. In addition, it was shown that the effectiveness of the surfactant could be weakened by wax molecules. Moreover, there existed a critical wax content above which the effectiveness of the surfactant was greatly reduced and the critical wax content gradually increased with increasing surfactant concentration. This work could provide guidance for hydrate management in wax-containing systems.