Biology subjects

Zhou, H.

Publications and source records attributed to Zhou, H..

At least 397 records · Page 22Linked to original sources

bioRxiv2020

Structural basis for the inhibition of the papain-like protease of SARS-CoV-2 by small molecules

SARS-CoV-2 is the pathogen responsible for the COVID-19 pandemic. The SARS-CoV-2 papain-like cysteine protease has been implicated in virus maturation, dysregulation of host inflammation and antiviral immune responses. We showed that PLpro preferably cleaves the K48-ubiquitin linkage while also being capable of cleaving ISG15 modification. The multiple functions of PLpro render it a promising drug target. Therefore, we screened an FDA-approved drug library and also examined available inhibitors against PLpro. Inhibitor GRL0617 showed a promising IC50 of 2.1 M. The co-crystal structure of SARS-CoV-2 PLpro-C111S in complex with GRL0617 suggests that GRL0617 is a non-covalent inhibitor. NMR data indicate that GRL0617 blocks the binding of ISG15 to PLpro. The antiviral activity of GRL0617 reveal that PLpro is a promising drug target for therapeutically treating COVID-19. One Sentence SummaryCo-crystal structure of PLpro in complex with GRL0617 reveals the druggability of PLpro for SARS-CoV-2 treatment.

biochemistry↗

bioRxiv2020

SARS-CoV-2 spike glycoprotein vaccine candidate NVX-CoV2373 elicits immunogenicity in baboons and protection in mice

The COVID-19 pandemic continues to spread throughout the world with an urgent need for a safe and protective vaccine to effectuate herd immunity to control the spread of SARS-CoV-2. Here, we report the development of a SARS-CoV-2 subunit vaccine (NVX-CoV2373) produced from the full-length spike (S) protein, stabilized in the prefusion conformation. Purified NVX-CoV2373 S form 27.2nm nanoparticles that are thermostable and bind with high affinity to the human angiotensin-converting enzyme 2 (hACE2) receptor. In mice and baboons, low-dose NVX-CoV2373 with saponin-based Matrix-M adjuvant elicits high titer anti-S IgG that is associated with blockade of hACE2 receptor binding, virus neutralization, and protection against SARS-CoV-2 challenge in mice with no evidence of vaccine-associated enhanced respiratory disease (VAERD). NVX-CoV2373 vaccine also elicits multifunctional CD4+ and CD8+ T cells, CD4+ T follicular helper T cells (Tfh), and the generation of antigen-specific germinal center (GC) B cells in the spleen. These results support the ongoing phase 1/2 clinical evaluation of the safety and immunogenicity of NVX-CoV2327 with Matrix-M (NCT04368988).

microbiology↗

bioRxiv2020

A Targeted Vaccine against COVID-19: S1-Fc Vaccine Targeting the Antigen-Presenting Cell Compartment Elicits Protection against SARS-CoV-2 Infection

Vaccination efficacy is enhanced by targeting the antigen-presenting cell compartment. Here, we show that S1-Fc antigen delivery targeting the Fc{gamma}R+ antigen-presenting cell compartment elicits anti-SARS-CoV-2 S1-antigen specific IgG production in vivo exerting biologically functional and protective activity against live virus infection, assessed in a stringent experimental virus challenge assay in vitro. The S1-domain of the SARS-CoV-2 spike protein was genetically fused to a human immunoglobulin Fc moiety, which contributes to mediate S1-Fc cellular internalization by Fc{gamma}R+ antigen-presenting cells. Immediately upon administration intramuscularly, our novel vaccine candidate recombinant rS1-Fc homes to lymph nodes in vivo where Fc{gamma}R+ antigen-presenting cells reside. Seroconversion is achieved as early as day 7, mounting considerably increased levels of anti-S1 IgGs in vivo. Interestingly, immunization at elevated doses with non-expiring S1-Fc encoding dsDNA favors the education of a desired antigen-specific adaptive T cell response. However, low-dose immunization, safeguarding patient safety, using recombinant rS1-Fc, elicits a considerably elevated protection amplitude against live SARS-CoV-2 infection. Our promising findings on rS1-Fc protein immunization prompted us to further develop an affordable and safe product for delivery to our communities in need for COVID-19 vaccinations.

microbiology↗

bioRxiv2020

SARS-CoV-2 envelope protein causes acute respiratory distress syndrome (ARDS)-like pathological damage and constitutes an antiviral target

Cytokine storm and multi-organ failure are the main causes of SARS-CoV-2-related death. However, the origin of the virus excessively damaging abilities remains unknown. Here we show that the SARS-CoV-2 envelope (2-E) protein alone is sufficient to cause acute respiratory distress syndrome (ARDS)-like damage in vitro and in vivo. Overexpression of 2-E protein induced rapid pyroptosis-like cell death in various susceptible cells and robust secretion of cytokines and chemokines in macrophages. Intravenous administration of purified 2-E protein into mice caused ARDS-like pathological damage in lung and spleen. Overexpressed 2-E protein formed cation channels in host cell membranes, eventually leading to membrane rupture. Newly identified channel inhibitors exhibited potent anti-SARS-CoV-2 activity and excellent protective effects against the 2-E-induced damage both in vitro and in vivo. Importantly, their channel inhibition, cell protection and antiviral activities were positively correlated with each other, supporting 2-E is a promising drug target against SARS-CoV-2.

microbiology↗

bioRxiv2020

Loss of endothelial glucocorticoid receptor accelerates diabetic nephropathy

Endothelial cells play a key role in the regulation of disease and other developmental processes. Defective regulation of endothelial cell homeostasis may cause mesenchymal activation of other endothelial cells by autocrine effects or of neighboring cell types by paracrine effects, and in both cases contribute to organ fibrosis. However, regulatory control of endothelial cell homeostasis, is not well studied. Diabetes induced renal fibrosis in endothelial GR knock out mice (GRfl/fl;Tie 1 Cre; GRECKO) but not in control mice (GRfl/fl); hypercholesterolemia further enhanced severe renal fibrosis in diabetic GRECKO; Apoe-/- (DKO) but not in diabetic littermates (GRfl/fl; Apoe-/-). The fibrogenic phenotype in the kidneys of diabetic GRECKO and diabetic DKO were associated with aberrant cytokine and chemokine reprogramming. Canonical Wnt signaling was identified as new target for the action of endothelial GR. Wnt inhibiton improved kidney fibrosis by mitigating endothelial-to-mesenchymal transition (EndMT) and epithelial-to-mesenchymal transitions (EMT). Similarly, activation of fatty acid oxidation also suppressed kidney fibrosis. Conditioned media from endothelial cells from diabetic GRECKO stimulated Wnt signaling-dependent epithelial-to-mesenchymal transition in tubular epithelial cells from diabetic controls. These data demonstrate that endothelial GR is an essential antifibrotic core molecule in diabetes.

cell biology↗

bioRxiv2020

Cerebellar nuclei evolved by repeatedly duplicating a conserved cell type set

How have complex brains evolved from simple circuits? Here we investigated brain region evolution at cell type resolution in the cerebellar nuclei (CN), the output structures of the cerebellum. Using single-nucleus RNA sequencing in mice, chickens, and humans, as well as STARmap spatial transcriptomic analysis and whole-CNS projection tracing in mice, we identified a conserved cell type set containing two classes of region-specific excitatory neurons and three classes of region-invariant inhibitory neurons. This set constitutes an archetypal CN that was repeatedly duplicated to form new regions. Interestingly, the excitatory cell class that preferentially funnels information to lateral frontal cortices in mice becomes predominant in the massively expanded human Lateral CN. Our data provide the first characterization of CN transcriptomic cell types in three species and suggest a model of brain region evolution by duplication and divergence of entire cell type sets.

neuroscience↗

bioRxiv2020

Chloroquine and hydroxychloroquine as ACE2 blockers to inhibit viropexis of COVID-19 Spike psedotype virus

BackgroundThe novel coronavirus disease (2019-nCoV) has been affecting global health since the end of 2019 and there is no sign that the epidemic is abating. The major issue for controlling the infectious is lacking efficient prevention and therapeutic approaches. Chloroquine (CQ) and Hydroxychloroquine (HCQ) have been reported to treat the disease, but the underlying mechanism remains controversial. PurposeThe objective of this study is to investigate whether CQ and HCQ could be ACE2 blockers and used to inhibit 2019-nCoV virus infection. MethodsIn our study, we used CCK-8 staining, flow cytometry and immunofluorescent staining to evaluate the toxicity and autophagy of CQ and HCQ, respectively, on ACE2 high-expressing HEK293T cells (ACE2h cells). We further analyzed the binding character of CQ and HCQ to ACE2 by molecular docking and surface plasmon resonance (SPR) assays, 2019-nCoV spike pseudotyped virus was also used to observe the viropexis effect of CQ and HCQ in ACE2h cells. ResultsResults showed that HCQ is slightly more toxic to ACE2h cells than CQ. Both CQ and HCQ could bind to ACE2 with KD =(7.31{+/-}0.62)e-7 M and (4.82{+/-}0.87)e-7 M, respectively. They exhibit equivalent suppression effect for the entrance of 2019-nCoV spike pseudotyped virus into ACE2h cells. ConclusionsCQ and HCQ both inhibit the entrance 2019-nCoV into cells by blocking the binding of the virus with ACE2. Our findings provide novel insights into the molecular mechanism of CQ and HCQ treatment effect on virus infection.

biochemistry↗

bioRxiv2020

Crystal structure of SARS-CoV-2 main protease in complex with a Chinese herb inhibitor shikonin

Main protease (Mpro, also known as 3CLpro) has a major role in the replication of coronavirus life cycle and is one of the most important drug targets for anticoronavirus agents. Here we report the crystal structure of main protease of SARS-CoV-2 bound to a previously identified Chinese herb inhibitor shikonin at 2.45 angstrom resolution. Although the structure revealed here shares similar overall structure with other published structures, there are several key differences which highlight potential features that could be exploited. The catalytic dyad His41-Cys145 undergoes dramatic conformational changes, and the structure reveals an unusual arrangement of oxyanion loop stabilized by the substrate. Binding to shikonin and binding of covalent inhibitors show different binding modes, suggesting a diversity in inhibitor binding. As we learn more about different binding modes and their structure-function relationships, it is probable that we can design more effective and specific drugs with high potency that can serve as effect SARS-CoV-2 anti-viral agents.

biophysics↗

bioRxiv2020

Super-resolution segmentation network for reconstruction of packed neurites

Neuron reconstruction can provide the quantitative data required for measuring the neuronal morphology and is crucial in the field of brain research. However, the difficulty in reconstructing packed neuritis, wherein massive labor is required for accurate reconstruction in most cases, has not been resolved. In this work, we provide a fundamental pathway for solving this challenge by proposing the use of the super-resolution segmentation network (SRSNet) that builds the mapping of the neurites in the original neuronal images and their segmentation in a higher-resolution space. SRSNet focuses on enlarging the distances between the boundaries of the packed neurites producing the high-resolution segmentation images. Thus, in the construction of the training datasets, only the traced skeletons of neurites are required, which vastly increase the usability of SRSNet. From the results of the experiments conducted in this work, it has been observed that SRSNet achieves accurate reconstruction of packed neurites where the other state-of-the-art methods fail.

neuroscience↗

bioRxiv2020

SUMO orchestrates multiple alternative DNA-protein crosslink repair pathways

Several endogenous metabolites, environmental agents, and therapeutic drugs promote formation of covalent DNA-protein crosslinks (DPCs). Persistent DPCs pose a serious threat to genome integrity and are eliminated by multiple repair pathways. Aberrant Top1 crosslinks to DNA, or Top1ccs, are processed by Tdp1 and Wss1 functioning in parallel pathways in Saccharomyces cerevisiae. It remains obscure how cells choose between these diverse mechanisms of DPC repair. Here we show that several SUMO biogenesis factors - Ulp1, Siz2, Slx5, Slx8 - control repair of Top1cc or an analogous DPC lesion. Genetic analysis reveals that SUMO promotes Top1cc processing in the absence of Tdp1 but has an inhibitory role if cells additionally lack Wss1. In the tdp1{Delta} wss1{Delta} mutant, the E3 SUMO ligase Siz2 stimulates sumoylation in the vicinity of the DPC, but not SUMO conjugation to Top1. This Siz2-dependent sumoylation delays DPC repair when cells progress through S and G2 phases. Our findings suggest that SUMO tunes available repair pathways to facilitate faithful DPC repair.

molecular biology↗

bioRxiv2020

ANANSE: An enhancer network-based computational approach for predicting key transcription factors in cell fate determination

Proper cell fate determination is largely orchestrated by complex gene regulatory networks centered around transcription factors. However, experimental elucidation of key transcription factors that drive cellular identity is currently often intractable. Here, we present ANANSE (ANalysis Algorithm for Networks Specified by Enhancers), a network-based method that exploits enhancer-encoded regulatory information to identify the key transcription factors in cell fate determination. As cell type-specific transcription factors predominantly bind to enhancers, we use regulatory networks based on enhancer properties to prioritize transcription factors. First, we predict genome-wide binding profiles of transcription factors in various cell types using enhancer activity and transcription factor binding motifs. Subsequently, applying these inferred binding profiles, we construct cell type-specific gene regulatory networks, and then predict key transcription factors controlling cell fate transitions using differential networks between cell types. This method outperforms existing approaches in correctly predicting major transcription factors previously identified to be sufficient for trans-differentiation. Finally, we apply ANANSE to define an atlas of key transcription factors in 18 normal human tissues. In conclusion, we present a ready-to-implement computational tool for efficient prediction of transcription factors in cell fate determination and to study transcription factor-mediated regulatory mechanisms. ANANSE is freely available at https://github.com/vanheeringen-lab/ANANSE.

systems biology↗

bioRxiv2020

ScRNA-seq discover cell cluster change under OAB: ACE2 expression reveal possible alternation of 2019-nCoV infectious pathway

ObjectivePrevious study indicated that bladder cells which express ACE2 were a potential infection route of 2019-nCov. This study observed some differences of bladder cell cluster and their ACE2 expression between OAB mice and healthy mice, indicating the change of infectious possibility and pathway under overactive bladder (OAB) circumstance. Material and methodPubic dataset acquisition was used to get ACE2 expression in normal human bladder and mice bladder (GSE129845). We built up over OAB model and studied the impact on cell typing and ACE2 expression. By way of using single-cell RNA sequencing (scRNA-seq) technique, bladder cell clustering and ACE2 expression in various cell types were measured respectively. ResultIn pubic database (healthy human and mice bladder), ACE2 expression in humans and mice is concentrated in bladder epithelial cells. The disappearance of umbrella cells, a component of bladder epithelial, was found in our OAB model. In the two mouse bladder samples, ACE2 expression of epithelial cells is 34.1%, also the highest of all cell types. ConclusionThe disappearance of umbrella cell may alternate the infection pathway of 2019-nCov and relate to the onset and progression of OAB.

cell biology↗

bioRxiv2020

Microbubble-Mediated Cavitation Promotes Apoptosis and Suppresses Invasion in AsPC-1 Cells

This study aimed to identify the potential and mechanisms of microbubble-mediated cavitation in promoting apoptosis and suppressing invasion in cancer cells. AsPC-1 cells were used and divided into four groups: control group, microbubble-only (MB) group, ultrasound-only (US) group, and ultrasound combined with microbubbles (US+MB) group. Pulse ultrasound was used with a frequency of 360 kHz and an intensity of 0.7 W/cm2 for 1 min (duty rate=50%). Then cells in four groups were cultured for 24 h. Cell counting kit-8 illustrated that US+MB could decrease cell viability. Western blot confirmed that US+MB increased cleaved Caspase-3, Bcl-2-associated X protein (BAX), and decreased the B-cell lymphoma-2 (Bcl-2) levels. Besides, US+MB increased intracellular calcium ions and down-regulated cleaved Caspase-8. For proliferation ability, cells in US+MB group had a lower expression of Ki67 and the weakened colony formation ability. The transwell invasion assay showed that US+MB could decrease invasion ability in AsPC-1 cells. Further evidence showed that cells conducted with US+MB had the lower level of hypoxia-inducible factor-1 (HIF-1) and Vimentin and the higher expression of E-cadherin than the other three groups. Finally, cells conducted with US+MB had less invadopodia formation. In conclusion, these results suggested that microbubble-mediated cavitation promoted apoptosis and suppressed invasion in AsPC-1 cells. SignificanceCancer cells have a high rate of cell division and a high rate of cell division can speed up the development of hypoxia which lead to promote cancer invasion and metastasis. Here we used microbubble-mediated cavitation to generate a mechanical shock wave which could effectively kill cancer cells through physical damage and intrinsic signaling pathway. Furthermore, effective killing of cancer cells could restrain the development of hypoxia and prevent invasion ability of cancer cells.

bioengineering↗

bioRxiv2020

The legacy of C4 evolution in the hydraulics of C3 and C4 grasses

The anatomical reorganization required for optimal C4 photosynthesis should also impact plant hydraulics. Most C4 plants possess large bundle-sheath cells and high vein density, which should also lead to higher leaf hydraulic conductance (Kleaf) and capacitance. Paradoxically, the C4 pathway reduces water demand and increases water-use-efficiency, creating a potential mis-match between supply capacity and demand in C4 plant water relations. We use phylogenetic analyses, physiological measurements, and models to examine the reorganization of hydraulics in closely-related C4 and C3 grasses. Evolutionarily young C4 lineages have higher Kleaf, capacitance, turgor-loss-point, and lower stomatal conductance than their C3 relatives. In contrast, species from older C4 lineages show decreased Kleaf and capacitance, indicating that over time, C4 plants have evolved to optimize hydraulic investments while maintaining C4 anatomical requirements. The initial "over-plumbing" of C4 plants disrupts the positive correlation between maximal assimilation rate and Kleaf, decoupling a key relationship between hydraulics and photosynthesis generally observed in vascular plants.

ecology↗

bioRxiv2020

Optimal reorganization of photosynthesis after the evolution of C4

C3 and C4 are major functional types in terrestrial biosphere models, with photosynthesis traits as important input parameters. The evolution of C4 required reorganizations of Calvin-Benson-cycle and coordination of C4-cycle enzymes, resulting in divergences of physiological traits between C3 and C4. In addition, photosynthesis further optimized after the evolution of C4 causing diversification within C4 lineages due to different evolutionary histories. We combined optimality modeling, physiological measurements and phylogenetic analysis to examine how various aspects of C4 photosynthetic machinery were reorganized and coordinated within C4 lineages and as compared to closely-related C3 in grasses. Optimality models and measurements indicated a higher maximal electron transport to maximal Rubisco carboxylation ratio (Jmax/Vcmax) in C4 than C3, consistent with the optimal prediction to maximize photosynthesis. The coordination between Calvin-Benson and C4 cycles (Vpmax/Vcmax), however, is in line with the optimal modeling results under 200 ppm, as opposed to current CO2. Such inconsistencies can be explained by a slowly declining assimilation rate beyond optimal Vpmax/Vcmax. Although rapid coordination occurred early in C4 evolution, C4 is still under optimizing processes and photosynthetic measures have continued to increase across time. Lastly, better understandings of Jmax/Vcmax, Vpmax/Vcmax and fluorescence-based-electron-transport proffer enhanced approaches to parameterize terrestrial biosphere models.

ecology↗

bioRxiv2020

VST Family Proteins are Regulators of Root System Architecture in Rice and Arabidopsis

Root System Architecture (RSA) is a key factor in the efficiency of nutrient capture and water uptake in plants. Understanding the genetic control of RSA will be useful in minimizing fertilizer and water usage in agricultural cropping systems. Using a hydroponic screen and a gel-based imaging system we identified a rice gene, OsVST1, which plays a key role in controlling RSA. This gene encodes a homolog of the Arabidopsis VAP-RELATED SUPPRESSORS OF TMM (VSTs), a class of proteins that promote signaling in stomata by mediating plasma membrane-endoplasmic reticulum contacts. OsVST1 mutants have shorter primary roots, decreased root meristem size, and a more compact root system architecture. We show that the Arabidopsis VST triple mutants have similar phenotypes, with reduced primary root growth and smaller root meristems. Expression of OsVST1 largely complements the short root length and reduced plant height in the Arabidopsis triple mutant, supporting conservation of function between rice and Arabidopsis VST proteins. In a field trial, mutations in OsVST1 do not adversely affect grain yield, suggesting that modulation of this gene could be used as a way to optimize RSA without an inherent yield penalty.

plant biology↗

bioRxiv2020

Structure of SARS-CoV-2 main protease in the apo state reveals the inactive conformation

Mpro is of considerable interest as a drug target in the treatment of COVID-19 since the proteolytic activity of this viral protease is essential for viral replication. Here we report the first insight of the structure Mpro for SARS-CoV-2 in the inactive conformation under conditions close to the physiological state (pH 7.5) to an overall resolution of 1.9 [A]. The comparisons of Mpro in different states reveal that substrate binding site and the active site are more flexible in the inactive conformation than that in the active conformations. Notably, compared with the active conformation of the apo state structure in pH7.6 of SARS, the SARS-CoV-2 apo state is in the inactive conformation under condition close to physiological state (pH7.5). Two water molecules are present in the oxyanion hole in our apo state structure, whereas in the ligand-bound structure, water molecular is absence in the same region. This structure provides novel and important insights that have broad implications for understanding the structural basis underlying enzyme activity, and can facilitate rational, structure-based, approaches for the design of specific SARS-CoV-2 ligands as new therapeutic agents.

biochemistry↗

bioRxiv2020

Dexmedetomidine attenuates the injury of H9C2 cardiomyocytes under Hypoxia/reoxygenation condition partly through the inhibition of endoplasmic reticulum stress

Background Myocardial ischemia-reperfusion injury (MIRI) has been confirmed to induce endoplasmic reticulum stress(ERS) during downstream cascade reaction when myocardial cell function keep deteriorating to a certain degree. The fact of matter is the clinical inconsistence with experimental outcomes still exist due to the mechanism has not been entirely clarified. Dexmedetomidine (DEX), a new generation anti-inflammatory and organ protector, has been testified can attenuate the IRI of heart. This study aimed to find out if DEX had the capacity to protect the injured cardiomyocytes under in vitro hypoxia/reoxygenation circumstance and if the ERS was totally or partly intervened.Methods H9C2 cells were subjected to cytotoxicity detection for 24h with DEX normally cultivated in several different concentrations. The proper hypoxia/reoxygenation (H/R) model parameter were concluded by the cell viability and injuries by cell counting kit-8(CCK8) and lactate dehydrogenase (LDH) release, when undergoing hypoxic condition for 3 h and reoxygenated for 3h, 6h,12h, and 24h, respectively. Also, the above index was assessed for H/R cardiomyocytes cultivated by various concentrations of DEX. The apoptosis, expression of) Glucose-regulated protein 78(GRP78), C/EBP homologous protein (CHOP), and caspase-12 were also examined in all groups.Results 1, 5 and 10 μM DEX in normal culture could significantly promote the proliferation of H9C2 (> 80%); the activity of H9c2 cells decreased to 62.67% (P < 0.05) at 3h of reoxygenation and to 36% at 6h of reoxygenation followed by 3h anoxic treatment; The cell viability of H9c2 cells in H/R groups incubated with 1 μM DEX increased 61.3%, and the LDH concentration in the supernatant was effectively lowered (−13.7, P < 0.05); H/R dramatically decreased the proportion of flow cytometry apoptosis and increased the expression of GRP78, CHOP and caspase-12, while both DEX and 4-phenyl butyric acid (4-PBA) could significantly reverse those above indicators. Additionally, DEX could induce deeper alterations than 4-PBA on the basis of H/R.Conclusion 1 μM DEX can dramatically attended the cell injuries, apoptosis, the expression of GRP78, CHOP and caspase-12 of H9C2 induced by 3h’ hypoxia and 3h’s reoxygenation. moreover, the functions of DEX went beyond the inhibition of ERS under this situation.View Full Text

scientific communication and education↗