A pseudovirus system enables deep mutational scanning of the full SARS-CoV-2 spike.

A major challenge in understanding SARS-CoV-2 evolution is interpreting the antigenic and functional effects of emerging mutations in the viral spike protein. Here, we describe a deep mutational scanning platform based on non-replicative pseudotyped lentiviruses that directly quantifies how large numbers of spike mutations impact antibody neutralization and pseudovirus infection. We apply this platform to produce libraries of the Omicron BA.1 and Delta spikes. These libraries each contain ∼7,000 distinct amino acid mutations in the context of up to ∼135,000 unique mutation combinations. We use these libraries to map escape mutations from neutralizing antibodies targeting the receptor-binding domain, N-terminal domain, and S2 subunit of spike. Overall, this work establishes a high-throughput and safe approach to measure how ∼105 combinations of mutations affect antibody neutralization and spike-mediated infection. Notably, the platform described here can be extended to the entry proteins of many other viruses.

Targeted isolation of diverse human protective broadly neutralizing antibodies against SARS-like viruses.

The emergence of current severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) and potential future spillovers of SARS-like coronaviruses into humans pose a major threat to human health and the global economy. Development of broadly effective coronavirus vaccines that can mitigate these threats is needed. Here, we utilized a targeted donor selection strategy to isolate a large panel of human broadly neutralizing antibodies (bnAbs) to sarbecoviruses. Many of these bnAbs are remarkably effective in neutralizing a diversity of sarbecoviruses and against most SARS-CoV-2 VOCs, including the Omicron variant. Neutralization breadth is achieved by bnAb binding to epitopes on a relatively conserved face of the receptor-binding domain (RBD). Consistent with targeting of conserved sites, select RBD bnAbs exhibited protective efficacy against diverse SARS-like coronaviruses in a prophylaxis challenge model in vivo. These bnAbs provide new opportunities and choices for next-generation antibody prophylactic and therapeutic applications and provide a molecular basis for effective design of pan-sarbecovirus vaccines.

Broadly neutralizing anti-S2 antibodies protect against all three human betacoronaviruses that cause deadly disease.

Pan-betacoronavirus neutralizing antibodies may hold the key to developing broadly protective vaccines against novel pandemic coronaviruses and to more effectively respond to SARS-CoV-2 variants. The emergence of Omicron and subvariants of SARS-CoV-2 illustrates the limitations of solely targeting the receptor-binding domain (RBD) of the spike (S) protein. Here, we isolated a large panel of broadly neutralizing antibodies (bnAbs) from SARS-CoV-2 recovered-vaccinated donors, which targets a conserved S2 region in the betacoronavirus spike fusion machinery. Select bnAbs showed broad in vivo protection against all three deadly betacoronaviruses, SARS-CoV-1, SARS-CoV-2, and MERS-CoV, which have spilled over into humans in the past two decades. Structural studies of these bnAbs delineated the molecular basis for their broad reactivity and revealed common antibody features targetable by broad vaccination strategies. These bnAbs provide new insights and opportunities for antibody-based interventions and for developing pan-betacoronavirus vaccines.

GSK3ß: A ray of hope for the treatment of diabetic kidney disease.

Diabetic kidney disease (DKD), a major microvascular complication of diabetes, is characterized by its complex pathogenesis, high risk of chronic renal failure, and lack of effective diagnosis and treatment methods. GSK3ß (glycogen synthase kinase 3ß), a highly conserved threonine/serine kinase, was found to activate glycogen synthase. As a key molecule of the glucose metabolism pathway, GSK3ß participates in a variety of cellular activities and plays a pivotal role in multiple diseases. However, these effects are not only mediated by affecting glucose metabolism. This review elaborates on the role of GSK3ß in DKD and its damage mechanism in different intrinsic renal cells. GSK3ß is also a biomarker indicating the progression of DKD. Finally, the protective effects of GSK3ß inhibitors on DKD are also discussed.

c-Cbl induced podocin ubiquitination contributes to the podocytes injury in diabetic nephropathy.

The ubiquitination function in diabetic nephropathy (DN) has attracted much attention, but there is a lack of information on its ubiquitylome profile. To examine the differences in protein content and ubiquitination in the kidney between db/db mice and db/m mice, we deployed liquid chromatography-mass spectrometry (LC-MS/MS) to conduct analysis. We determined 145 sites in 86 upregulated modified proteins and 66 sites in 49 downregulated modified proteins at the ubiquitinated level. Moreover, 347 sites among the 319 modified proteins were present only in the db/db mouse kidneys, while 213 sites among the 199 modified proteins were present only in the db/m mouse kidneys. The subcellular localization study indicated that the cytoplasm had the highest proportion of ubiquitinated proteins (31.87%), followed by the nucleus (30.24%) and the plasma membrane (20.33%). The enrichment analysis revealed that the ubiquitinated proteins are mostly linked to tight junctions, oxidative phosphorylation, and thermogenesis. Podocin, as a typical protein of slit diaphragm, whose loss is a crucial cause of proteinuria in DN. Consistent with the results of ubiquitination omics, the K261R mutant of podocin induced the weakest ubiquitination compared with the K301R and K370R mutants. As an E3 ligase, c-Cbl binds to podocin, and the regulation of c-Cbl can impact the ubiquitination of podocin. In conclusion, in DN, podocin ubiquitination contributes to podocyte injury, and K261R is the most significant site. c-Cbl participates in podocin ubiquitination and may be a direct target for preserving the integrity of the slit diaphragm structure, hence reducing proteinuria in DN.

Electron pairing in the pseudogap state revealed by shot noise in copper oxide junctions.

In the quest to understand high-temperature superconductivity in copper oxides, debate has been focused on the pseudogap-a partial energy gap that opens over portions of the Fermi surface in the 'normal' state above the bulk critical temperature1. The pseudogap has been attributed to precursor superconductivity, to the existence of preformed pairs and to competing orders such as charge-density waves1-4. A direct determination of the charge of carriers as a function of temperature and bias could help resolve among these alternatives. Here we report measurements of the shot noise of tunnelling current in high-quality La2-xSrxCuO4/La2CuO4/La2-xSrxCuO4 (LSCO/LCO/LSCO) heterostructures fabricated using atomic layer-by-layer molecular beam epitaxy at several doping levels. The data delineate three distinct regions in the bias voltage-temperature space. Well outside the superconducting gap region, the shot noise agrees quantitatively with independent tunnelling of individual charge carriers. Deep within the superconducting gap, shot noise is greatly enhanced, reminiscent of multiple Andreev reflections5-7. Above the critical temperature and extending to biases much larger than the superconducting gap, there is a broad region in which the noise substantially exceeds theoretical expectations for single-charge tunnelling, indicating pairing of charge carriers. These pairs are detectable deep into the pseudogap region of temperature and bias. The presence of these pairs constrains current models of the pseudogap and broken symmetry states, while phase fluctuations limit the domain of superconductivity.

A broad and potent neutralization epitope in SARS-related coronaviruses.

Many neutralizing antibodies (nAbs) elicited to ancestral severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) through natural infection and vaccination have reduced effectiveness to SARS-CoV-2 variants. Here, we show that therapeutic antibody ADG20 is able to neutralize SARS-CoV-2 variants of concern (VOCs) including Omicron (B.1.1.529) as well as other SARS-related coronaviruses. We delineate the structural basis of this relatively escape-resistant epitope that extends from one end of the receptor binding site (RBS) into the highly conserved CR3022 site. ADG20 can then benefit from high potency through direct competition with ACE2 in the more variable RBS and interaction with the more highly conserved CR3022 site. Importantly, antibodies that are able to target this site generally neutralize a broad range of VOCs, albeit with reduced potency against Omicron. Thus, this conserved and vulnerable site can be exploited for the design of universal vaccines and therapeutic antibodies.

Total Synthesis of (±)-Rubriflordilactone A.

A new and efficient synthesis of rubriflordilactone A has been realized. The key transformations include the following: (1) an intramolecular Prins cyclization to establish the seven-membered ring containing two contiguous stereocenters; (2) a Mukaiyama hydration/oxa-Michael cascade to construct the B-ring; and (3) an unprecedented stereocontrol intermolecular o-QM type [4 + 2]-cycloaddition to rapidly assemble core structure of rubriflordilactone A.

Mechanism and origin of enantioselectivity for organocatalyzed asymmetric heteroannulation of alkynes in the construction of axially chiral C2-arylquinoline.

Axially chiral C2-arylquinoline has been successfully constructed via asymmetric heteroannulation of alkynes catalyzed by chiral phosphoric acid with high yield and high enantioselectivity. Inspired by this intriguing work, theoretical calculations have been carried out, and the detailed reaction mechanism has been elaborated, in which the whole reaction can be divided into steps including hydrogen transfer, C-N bonding, annulation reaction and the final dehydration processes. The initial hydrogen-transfer reaction has two possible pathways, while the subsequent C-N bonding process has eight possible pathways. Then, after the annulation reaction and the final dehydration processes, the major product and byproduct were formed. QTAIM and IGMH analyses were used to illustrate the role of weak intermolecular interactions in the catalytic process, and the distortion/interaction and EDA analyses provided a deeper understanding of the origin of enantioselectivity. The calculated results are consistent with the experimental results. This work would provide valuable insights into asymmetric reactions catalyzed by chiral phosphoric acid.

Changes of vertical distribution of Microcystis colonies driven by short-term rainfall: Disappearance and reformation of surface bloom.

Global climate warming and human activities have increased the magnitude and frequency of Microcystis surface blooms, posing significant threats to freshwater ecosystems and human health over recent decades. Heavy rainfall events have been reported to cause the disappearance of these blooms. Although some studies have employed turbulence models to analyze the movement characteristics of Microcystis colonies, the impact of rainfall is complex, comprehensive investigations on their vertical migration induced by short-term rainfall are still necessary. Utilizing monitoring data from eutrophic ponds and controlled simulation experiments, this study examines the short-term impacts of rainfall on the vertical distribution of Microcystis in the water column. Our findings indicate that rainfall contributes to the disappearance of Microcystis blooms by reducing the quantity of small to medium-sized colonies (0-100 µm) at the surface, subsequently decreasing the overall Microcystis biomass. As rainfall intensity increases, larger colonies migrate deeper into the water column. At a rainfall threshold of 666 mm, the difference in the median volume diameter (DV50) of Microcystis colonies between the surface and bottom reaches a minimal value of 3.09%. Post-rainfall, these colonies rapidly ascend, aggregate into larger formations, and re-establish surface blooms. The greater the rainfall, the smaller the resultant Microcystis biomass, albeit with larger aggregated colony sizes. When rainfall exceeds 222 mm, the recovery rate of surface Microcystis biomass remains below 100%, decreasing to 19.48% at 666 mm of rainfall, while the median volume diameter (DV50) of the colonies increases to 139.07% of its pre-rainfall level. Furthermore, compared to pre-rainfall conditions, the photosynthetic activity of the surface Microcystis colonies was enhanced and the secretion of EPS was increased under heavy rainfall conditions. Our results identify a critical response time of 30 min for Microcystis colonies to rainfall, after which the response ceases to intensify. These insights are crucial for predicting post-rain Microcystis bloom dynamics and aiding management authorities in timely interventions.

Enantiocontrolled Total Synthesis of (-)-Retigeranic Acid A.

The asymmetric total synthesis of (-)-retigeranic acid A has been realized. The key features of the current synthesis include (1) a Pt-catalyzed Conia-ene 5-exo-dig cyclization of enolyne to establish the key quaternary stereochemical center of C-10 (D/E ring), (2) an intramolecular diastereoselective Prins cyclization to construct the trans-hydrindane backbone (A/B ring), and (3) a late-stage intramolecular Fe-mediated hydrogen atom transfer (HAT) Baldwin-disfavored 5-endo-trig radical cyclization to rapidly assemble vicinal quaternary centers and the core structure of (-)-retigeranic acid A (C ring).

Fluorescent/Colorimetric Dual-Mode Discriminating Gln and Val Enantiomers Based on Carbon Dots.

Discrimination and quantification of amino acid (AA) enantiomers are particularly important for diagnosing and treating diseases. Recently, dual-mode probes have gained a lot of research interest because they can catch more detecting information compared with the single-mode probes. Thus, it is of great significance to develop a dual-mode sensor realizing AA enantiomer discrimination conveniently and efficiently. In this work, carbon dot L-TCDs were prepared by N-methyl-1,2-benzenediamine dihydrochloride (OTD) and l-tryptophan. With the assistance of H2O2, L-TCDs show an excellent discrimination performance for enantiomers of glutamine (Gln) and valine (Val) in both fluorescent and colorimetric modes. The fluorescence enantioselectivity of Gln (FD/FL) and Val (FL/FD) is 5.29 and 4.13, respectively, and the colorimetric enantioselectivity of Gln (ID/IL) and Val (IL/ID) is 13.26 and 3.42, individually. The chiral recognition mechanism of L-TCDs was systematically studied. L-TCDs can be etched by H2O2, and the participation of AA enantiomers results in different amounts of the released OTD, which provides fluorescent and colorimetric signals for identifying and quantifying the enantiomers of Gln and Val. This work provides a more convenient and flexible dual-mode sensing strategy for discriminating AA enantiomers, which is expected to be of great value in facile and high-throughput chiral recognition.

Network Analysis Indicates Microbial Assemblage Differences in Life Stages of Cladophora.

Cladophora represents a microscopic forest that provides many ecological niches and fosters a diverse microbiota. However, the microbial community on Cladophora in brackish lakes is still poorly understood. In this study, the epiphytic bacterial communities of Cladophora in Qinghai Lake were investigated at three life stages (attached, floating, and decomposing). We found that in the attached stage, Cladophora was enriched with chemoheterotrophic and aerobic microorganisms, including Yoonia-Loktanella and Granulosicoccus. The proportion of phototrophic bacteria was higher in the floating stage, especially Cyanobacteria. The decomposing stage fostered an abundance of bacteria that showed vertical heterogeneity from the surface to the bottom. The surface layer of Cladophora contained mainly stress-tolerant chemoheterotrophic and photoheterotrophic bacteria, including Porphyrobacter and Nonlabens. The microbial community in the middle layer was similar to that of floating-stage Cladophora. Purple oxidizing bacteria were enriched in the bottom layer, with Candidatus Chloroploca, Allochromatium, and Thiocapsa as the dominant genera. The Shannon and Chao1 indices of epibiotic bacterial communities increased monotonically from the attached stage to the decomposing stage. Microbial community composition and functional predictions indicate that a large number of sulfur cycle-associated bacteria play an important role in the development of Cladophora. These results suggest that the microbial assemblage on Cladophora in a brackish lake is complex and contributes to the cycling of materials. IMPORTANCE Cladophora represents a microscopic forest that provides many ecological niches fostering a diverse microbiota, with a complex and intimate relationship between Cladophora and bacteria. Many studies have focused on the microbiology of freshwater Cladophora, but the composition and succession of microorganisms in different life stages of Cladophora, especially in brackish water, have not been explored. In this study, we investigated the microbial assemblages in the life stages of Cladophora in the brackish Qinghai Lake. We show that heterotrophic and photosynthetic autotrophic bacteria are enriched in attached and floating Cladophora, respectively, whereas the epiphytic bacterial community shows vertical heterogeneity in decomposing mats.

Evaluation of the Curative Effect of Treating HBV-related Liver Fibrosis/Cirrhosis Based on the Method of Removing Turbidity and Regulating the Liver.

Objective: This study investigated the effectiveness of a technique for eliminating cloudiness and managing liver function in treating liver fibrosis/cirrhosis associated with the Hepatitis B virus (HBV). Methods: From January 2022 to January 2023, the researchers' hospital treated 200 patients with HBV-related liver fibrosis/cirrhosis. These patients constituted two groups for the study: the control group, consisting of 100 cases who received routine treatment, and a study group, consisting of 56 cases who received treatment with a combination of turbidity removal and liver regulation, in addition to the standard treatment given to the control group. The researchers then compared factors such as liver function, level of liver fibrosis, liver stiffness measurement (LSM), and renal function between the two groups. Additionally, the researchers assessed the effectiveness of those treatments and any adverse reactions that may have occurred. Results: The study group demonstrated significantly higher clinical effectiveness than the control group after undergoing treatment, with statistical significance (P < .05). Post-treatment, both groups experienced lower GGT, ALT, and AST levels than their pre-treatment levels. Additionally, the study group had higher AIB levels than their pre-treatment levels. There was a statistically significant difference between the study and control groups regarding these biomarkers (P < .05), as the study group exhibited lower GGT, ALT, AST, TBIL levels and higher AIB levels. Furthermore, both groups displayed decreased HA, IV-C, PC III, and LN levels post-treatment compared to their pre-treatment values. The study group had significantly lower HA, IV-C, PC III, and LN concentrations than the control group (P < .05). Regarding LSM measurements after treatment for both groups, while there was a decrease in LSM values from their respective pre-treatment readings for each group, no significant difference was observed between them (P < .05). Moreover, the incidence of adverse reactions experienced by individuals in the study group following treatment was significantly lower than that of individuals in the control group (P < .05). Conclusion: Treatment based on removing turbidity and regulating the liver can effectively relieve the clinical symptoms of patients with HBV-related liver fibrosis/cirrhosis, promote the liver function to return to normal, relieve the degree of liver fibrosis, and reduce the LSM value. The curative effect is significant and worthy of clinical application.

PVP-capped silver nanoparticles for efficient SERS detection of adenine based on the stabilizing and enrichment roles of PVP.

A polyvinylpyrrolidone-capped (PVP-capped) strategy is reported to synthesize Ag NPs on silicon wafers via galvanic replacement reaction for SERS detection of adenine, where PVP acts as stabilizing agent in synthesis and efficient enrichment in detection. The morphologies of Ag NPs are optimized with uniform particle size by adjusting synthesis conditions, which hold excellent SERS performances like a high enhancement factor of 1.42 × 106, good uniform, reproducibility, and transferable nature. With the protection of the capped PVP, the Ag NPs keep excellent SERS properties even against harsh conditions of high temperature (100 ℃) and strong acid and base for 24 h. Utilizing the structural feature of PVP with abundant carbonyl groups, the PVP-capped Ag NPs achieve efficient enrichment of adenine through hydrogen bonding and π-interactions, which is analyzed by density functional theory. Quantitative detection of adenine is performed with a wide linear range from 10-4 to 10-8 M and a low limit of detection of 1 nM. Detection of adenine in human urine samples is achieved with a recovery of 99.1-103.4% and an RSD of less than 5%.

The effects of intrauterine growth on physical and intellectual development of one-year-old infants: a study on monochorionic twins with selective intrauterine growth restriction.

This study aimed to evaluate physical and intellectual development of one-year-old infants of monochorionic twins with selective intrauterine growth restriction (sIUGR). A total of 31 pairs of sIUGR twins ageing 1 year old were included in the study. Each pair of sIUGR twins was divided into low birthweight-twin group (L-twin group) and high birthweight-twin group (H-twin group) according to twins' birthweight. The differences in height, weight, head circumstance and body mass index (BMI) in each stage were statistically significant for all measures from birth until 1 year old (p < .05), and there was a disappointed catch-up growth in lighter twins. Psychomotor development index (PDI) and mental development index (MDI) at 1 year old were significantly different between the two groups (p < .05). Stepwise regression analysis showed that the effects of weight on both PDI and MDI were statistically significant (p < .05). Intrauterine growth inconsistencies in monochorionic twins with sIUGR persist until the first year of life and affect low-birthweight infants' physical and intellectual development.Impact StatementWhat is already known on this subject? Selective intrauterine growth restriction in monochorionic twins increases the risks of intrauterine foetal demise, preterm birth, caesarean delivery and adverse neonatal outcomes, especially in the smaller foetus.What do the results of this study add? Previous studies have concentrated on the clinical management of sIUGR, while little attention has been paid to the growth and development of twins after birth. Given the adverse neurobiological effects of suboptimal nutrition on the brain development, it is important to determine whether IUGR causes long-term cognitive deficits and physical retardation. The current study has assessed the physical and intellectual development of one-year-old infants of monochorionic twins with sIUGR.What are the implications of these findings for clinical practice and/or further research? Intrauterine growth inconsistencies in monochorionic twins with sIUGR persist until the first year of life and affect low-birthweight infants' physical and intellectual development. Further research on the longer-term effects of sIUGR is needed.

Zr(OH)4 -Catalyzed Controllable Selective Oxidation of Anilines to Azoxybenzenes, Azobenzenes and Nitrosobenzenes.

The selective oxidation of aniline to metastable and valuable azoxybenzene, azobenzene or nitrosobenzene has important practical significance in organic synthesis. However, uncontrollable selectivity and laborious synthesis of the expensive required catalysts severely hinders the uptake of these reactions in industrial settings. Herein, we have pioneered the discovery of Zr(OH)4 as an efficient heterogeneous catalyst capable of the selective oxidation of aniline, using either peroxide or O2 as oxidant, to selectively obtain various azoxybenzenes, symmetric/unsymmetric azobenzenes, as well as nitrosobenzenes, by simply regulating the reaction solvent, without the need for additives. Mechanistic experiments and DFT calculations demonstrate that the activation of H2 O2 and O2 is primarily achieved by the bridging hydroxyl and terminal hydroxyl groups of Zr(OH)4 , respectively. The present work provides an economical and environmentally friendly strategy for the selective oxidation of aniline in industrial applications.

Broadly resistant HIV-1 against CD4-binding site neutralizing antibodies.

Recently identified broadly neutralizing antibodies (bnAbs) show great potential for clinical interventions against HIV-1 infection. However, resistant strains may impose substantial challenges. Here, we report on the identification and characterization of a panel of HIV-1 strains with broad and potent resistance against a large number of bnAbs, particularly those targeting the CD4-binding site (CD4bs). Site-directed mutagenesis revealed that several key epitope mutations facilitate resistance and are located in the inner domain, loop D, and ß23/loop V5/ß24 of HIV-1 gp120. The resistance is largely correlated with binding affinity of antibodies to the envelope trimers expressed on the cell surface. Our results therefore demonstrate the existence of broadly resistant HIV-1 strains against CD4bs neutralizing antibodies. Treatment strategies based on the CD4bs bnAbs must overcome such resistance to achieve optimal clinical outcomes.

Indole Alkaloids from a Soil-Derived Clonostachys rosea.

Clonorosins A (1) and B (2), two novel indole alkaloids featuring unprecedented 6/5/6/6/5 and 6/5/5 cores, together with seven known indole-linked 2,5-diketopiperazine alkaloids (3-9), were isolated from the soil-derived fungus Clonostachys rosea YRS-06. The new structures were proposed through HR-MS, NMR, and ECD spectroscopic data. They were established by comparing the calculated NMR, ECD, and specific rotation data with the experimental. To assist in determining the absolute configuration of the chiral carbon in the side chain of 2,5-diketopiperazine derivatives, flexible analogues 3i-3iv were synthesized and analyzed. 1 was active against Fusarium oxysporum with an MIC value of 50 µg/mL. 7 and 8 showed excellent activity against human HeLa and HepG2 cells with IC50 values of 0.12-0.60 µM.

Chalcogen⋠⋠⋠π Bonding Catalysis.

While the presence of sulfur⋅⋅⋅π bonding interaction is a general phenomenon in the biological systems, the exploitation of this noncovalent force in a chemical process yet remains elusive. Herein, we describe the concept of chalcogen⋅⋅⋅π bonding catalysis that activates molecules of π systems through the interaction between chalcogen and π-electron cloud. The proof-of-concept studies using a vinylindole-based Diels-Alder benchmark reaction demonstrate that S⋅⋅⋅π and Se⋅⋅⋅π bonding interaction can drive the cycloaddition reaction efficiently. Experimental results suggest that a simultaneously double Se⋅⋅⋅π bonding interaction directs the stereoselectivity in this cycloaddition process.