Biology subjects

Huang, W.

Publications and source records attributed to Huang, W..

At least 307 records · Page 17Linked to original sources

bioRxiv2020

Bridging the gap between in silico and in vivo: modeling opioid disposition in a kidney proximal tubule microphysiological system

BackgroundOpioid overdose, dependence, and addiction are a major public health crisis. Patients with chronic kidney disease (CKD) are at high risk of opioid overdose, therefore novel methods that provide accurate prediction of kidney clearance (CLr) and systemic disposition of opioids in CKD patients can facilitate the optimization of therapeutic regimens. MethodsWe conducted prediction of kidney clearance and systemic disposition of morphine and its active metabolite morphine-6-glucuronide (M6G) in CKD patients using a vascularized human proximal tubule microphysiological system (VPT-MPS) coupled with a parent-metabolite full body physiologically-based pharmacokinetic (PBPK) model. ResultsThe VPT-MPS, populated with a human umbilical vein endothelial cell (HUVEC) channel and an adjacent human primary proximal tubular epithelial cells (PTEC) channel, successfully demonstrated secretory transport of morphine and M6G from the HUVEC channel into the PTEC channel in a time-dependent manner; transporter inhibitors decreased translocation by 74.3% and 63.6%, respectively. The in vitro data generated by VPT-MPS were incorporated into a mechanistic kidney model and parent-metabolite full body PBPK model to predict CLr and systemic disposition of morphine and M6G. The model successfully predicted CLr within 1.5-fold, and the plasma concentration-time profiles of morphine and M6G in both healthy subjects and CKD patients, with absolute average fold error values <1.5. ConclusionsA microphysiological system together with mathematical modeling successfully predicted kidney clearance and systemic disposition of opioids in CKD patients and healthy subjects.

pharmacology and toxicology↗

bioRxiv2020

Double Lock of a Potent Human Monoclonal Antibody against SARS-CoV-2

Receptor recognition and subsequent membrane fusion are essential for the establishment of successful infection by SARS-CoV-2. Halting these steps can cure COVID-19. Here we have identified and characterized a potent human monoclonal antibody, HB27, that blocks SARS-CoV-2 attachment to its cellular receptor at sub-nM concentrations. Remarkably, HB27 can also prevent SARS-CoV-2 membrane fusion. Consequently, a single dose of HB27 conferred effective protection against SARS-CoV-2 in two established mouse models. Rhesus macaques showed no obvious adverse events when administrated with 10-fold of effective dose of HB27. Cryo-EM studies on complex of SARS-CoV-2 trimeric S with HB27 Fab reveal that three Fab fragments work synergistically to occlude SARS-CoV-2 from binding to ACE2 receptor. Binding of the antibody also restrains any further conformational changes of the RBD, possibly interfering with progression from the prefusion to the postfusion stage. These results suggest that HB27 is a promising candidate for immuno-therapies against COVID-19. HighlightsO_LISARS-CoV-2 specific antibody, HB27, blocks viral receptor binding and membrane fusion C_LIO_LIHB27 confers prophylactic and therapeutic protection against SARS-CoV-2 in mice models C_LIO_LIRhesus macaques showed no adverse side effects when administered with HB27 C_LIO_LICryo-EM studies suggest that HB27 sterically occludes SARS-CoV-2 from its receptor C_LI

biophysics↗

bioRxiv2020

An investigation of MHC-dependent sexual selection in a free-living population of sheep

The MHC is one of the most polymorphic gene clusters in vertebrates and play an essential role in adaptive immunity. Apart from pathogen-mediated selection, sexual selection can also contribute to the maintenance of MHC diversity. MHC-dependent sexual selection could occur via several mechanisms but at present there is no consensus as to which of these mechanisms are involved and their importance. Previous studies have often suffered from limited genetic and behavioural data and small sample size, and were rarely able to examine all the mechanisms together, determine whether signatures of MHC-based non-random mating are independent of genomic effects or differentiate whether MHC-dependent sexual selection takes place at the pre- or post-copulatory stage. In this study, we use Monte Carlo simulation to investigate evidence for non-random MHC-dependent mating patterns by all three mechanisms in a free-living population of Soay sheep. Using 1710 sheep diplotyped at the MHC class IIa region and genome-wide SNPs, together with field observations of consorts, we found sexual selection against a particular haplotype in males at the pre-copulatory stage and sexual selection against female MHC heterozygosity during the rut. We also found MHC-dependent disassortative mating at the post-copulatory stage, along with strong evidence of inbreeding avoidance at both stages. However, results from generalized linear mixed models suggest that the pattern of MHC-dependent disassortative mating could be a by-product of inbreeding avoidance. Our results therefore suggest that while multiple apparent mechanisms of non-random mating with respect to the MHC may occur, some of them have alternative explanations.

evolutionary biology↗

bioRxiv2020

Microfluidic guillotine reveals multiple timescales and mechanical modes of wound response in Stentor coeruleus

BackgroundWound healing is one of the defining features of life and is seen not only in tissues but also within individual cells. Understanding wound response at the single-cell level is critical for determining fundamental cellular functions needed for cell repair and survival. This understanding could also enable the engineering of single-cell wound repair strategies in emerging synthetic cell research. One approach is to examine and adapt self-repair mechanisms from a living system that already demonstrates robust capacity to heal from large wounds. Towards this end, Stentor coeruleus, a single-celled free-living ciliate protozoan, is a unique model because of its robust wound healing capacity. This capacity allows one to perturb the wounding conditions and measure their effect on the repair process without immediately causing cell death, thereby providing a robust platform for probing the self-repair mechanism. ResultsHere we used a microfluidic guillotine and a fluorescence-based assay to probe the timescales and mechanisms of wound repair in Stentor. We found that Stentor requires [~]100 - 1000 s to close bisection wounds, depending on the severity of the wound. This corresponds to a healing rate of [~]8 - 80 m2/s, faster than most other single cells reported in the literature. Further, we observed and characterized three distinct mechanical modes of wound repair in Stentor: contraction, cytoplasm retrieval, and twisting/pulling. Using chemical perturbations, active cilia were found to be important for only the twisting/pulling mode. Contraction of myonemes, a major contractile fiber in Stentor, was surprisingly not important for the contraction mode and was of low importance for the others. ConclusionsWhile events local to the wound site have been the focus of many single-cell wound repair studies, our results suggest that large-scale mechanical behaviors may be of greater importance to single-cell wound repair than previously thought. The work here advances our understanding of the wound response in Stentor, and will lay the foundation for further investigations into the underlying components and molecular mechanisms involved.

cell biology↗

bioRxiv2020

The Biogeography of Fungal Communities Across Different Chinese Wine-producing Regions Associated with Environmental Factors and Spontaneous Fermentation Performance

Chinese Marselan grapes are believed to possess the potential to become a characteristic regional variety, whose quality is internationally recognized. The fermentation-related mycobiota from six climatically diverse Marselan-producing regions in China were analyzed via high-throughput sequencing (HTS), while the influence of environmental factors was evaluated as well. The results implied that the phyla Ascomycota and genus Aureobasidium dominated the fungal communities in 166 Marselan must and ferment samples. Significant differences were detected in the fungal microbiota from the regions, as well as the wineries, while these discrepancies decreased as the fermentation progressed. Moreover, the difference in fungal communities between the wineries exceeded the variation involving the regions, and higher diversity was identified in the types of species than in the compositions. Geoclimatic elements (Gc) and physicochemical indexes (Pi) exerted a significant effect on the fungal must consortium, explaining 58.17% of the taxonomic information. Therefore, a correlation was proposed between the spontaneous fermentation performance, as well as the polyphenol (PP) content, and their association with fungal taxonomic composition. In addition to enriching the knowledge regarding the wine microbiome and its influencing factors, this study may provide a new strategy for harnessing autochthonous "microbial terroir". ImportanceWine microbiome and their metabolism play a crucial role in wine quality. Chinese wine-producing regions are widely distributed with diverse geoclimatic conditions, this study reports a most comprehensive biogeography of fermentation-related fungal communities performed to date, taking the Chinese promising Marselan grape variety as the research object. It reveals significant differences in the fungal microbiota of Marselan must and ferment from different regions and wineries, and higher otherness between species type than that between species composition. The study also reports the main geoclimatic and physicochemical factors shaping fungal flora. Moreover, a correlation was proposed among the spontaneous fermentation performance with fungal taxonomic composition and polyphenol content. Taken together, the results shed light on the wine fungal communities and its influencing factors, which is crucial for vineyard and fermentation microorganism management, and may also provide a new strategy for harnessing autochthonous "microbial terroir" by utilizing spontaneous fermentation.

microbiology↗

bioRxiv2020

Identification of a peroxidase inhibitor that enhances kanamycin activity

BackgroundThe threat of antimicrobial resistance calls for more efforts in basic science, drug discovery, and clinical development, particularly gram-negative carbapenem-resistant pathogens. Objectives and methodsWhole-cell-based screening was performed to identify novel antibacterial agents against Acinetobacter baumannii ATCC19606. Spontaneously resistant mutant selection, whole-genome sequencing, and surface plasmon resonance were used for target identification and confirmation. Checkerboard titration assay was used for drug combination analysis. ResultsA small molecule named 6D1 with the chemical structure of 6-fluorobenzo[d]isothiazol-3(2H)-one was identified and exhibited activity against A. baumannii ATCC19606 strain (minimal inhibitory concentration, MIC = 1 mg/L). The mutation in the plasmid-derived ohrB gene that encodes a peroxidase was identified in spontaneously resistant mutants. Treatment of the bacteria with 6D1 resulted in increased sensitivity to peroxide such as tert-butyl hydroperoxide. The binding of 6D1 and OhrB was confirmed by surface plasmon resonance. Interestingly, the MIC of kanamycin against spontaneously resistant mutants decreased. Finally, we identified the effect of 6D1 on enhancing the antibacterial activity of kanamycin, including New Delhi metallo-{beta}-lactamase (NDM-1)-producing carbapenem-resistant Klebsiella pneumoniae, but not in strains carrying kanamycin resistance genes. ConclusionsIn this study, we identified a peroxidase inhibitor that suppresses the growth of A. baumannii and enhances the antibacterial activity of kanamycin. We propose that peroxidase may be potentially used as a target for kanamycin adjuvant development.

microbiology↗

bioRxiv2020

Neuroinflammation characterizes the earliest changes in Alzheimer's disease pathology and associated subjective cognitive impairment in adult hydrocephalus biopsies

In an effort to better characterize the transcriptomic changes that accompany early Alzheimers disease (AD) pathology in living patients and correlate with contemporaneous cognitive data, we performed RNA-seq on 106 cortical biopsies that were taken during shunt placement for adult onset hydrocephalus with varying degrees of comorbid AD pathology. A restricted set of genes correlate with AD pathology in these biopsies, and co-expression network analysis demonstrates an evolution from microglial homeostasis to a disease-associated microglial phenotype in conjunction with increasing AD pathologic burden, along with a subset of additional astrocytic and neuronal genes that accompany these changes. Further analysis demonstrates that these correlations are driven by patients that report mild cognitive symptoms, despite similar levels of {beta}-amyloid and tau pathology in comparison to patients who report no cognitive symptoms. Interestingly, downregulation of homeostatic genes and upregulation of disease-associated genes also correlate with microglial plaque invasion and an activated microglial morphology, and this change is not sensitive to cognitive status, suggesting that an initial microglial response to AD pathology is eventually maladaptive. Taken together, these findings highlight a restricted set of microglial and non-microglial genes and suggest that early AD pathology is largely characterized by a loss of homeostatic genes and an activated microglial phenotype that continues to evolve in conjunction with accumulating AD pathology in the setting of subjective cognitive decline.

neuroscience↗

bioRxiv2020

The nuclear receptor RORα preserves cardiomyocyte mitochondrial function by regulating mitophagy through caveolin-3

Preserving optimal mitochondrial function is critical in the heart, which is the most ATP-avid organ in the body. Recently, we showed that global deficiency of the nuclear receptor ROR in the "staggerer" (RORsg/sg) mouse exacerbates angiotensin II-induced cardiac hypertrophy and compromises cardiomyocyte mitochondrial function. The mechanisms underlying these observations have not been defined. Here we present evidence that ROR regulates cardiomyocyte mitophagy using pharmacological and genetic gain- and loss-of-function tools, including a novel cardiomyocyte-specific ROR knockout mouse. Cardiomyocyte ROR is upregulated by hypoxia and the loss of ROR blunts hypoxia-induced mitophagy and broadly compromises mitochondrial function. We show that ROR is a direct transcriptional regulator of the mitophagy mediator caveolin-3 in cardiomyocytes and that increased expression of ROR increases caveolin-3 abundance and enhances mitophagy. Knockdown of ROR impairs cardiomyocyte mitophagy, but this defect can be rescued by caveolin-3 overexpression. Collectively, these findings reveal a novel role for ROR in regulating mitophagy through caveolin-3 and expand our currently limited understanding of the mechanisms underlying ROR-mediated cardioprotection.

molecular biology↗

bioRxiv2020

S-Trimer, a COVID-19 subunit vaccine candidate, induces protective immunity in nonhuman primates

SARS-CoV-2 is the underlying cause for the COVID-19 pandemic. Like most enveloped RNA viruses, SARS-CoV-2 uses a homotrimeric surface antigen to gain entry into host cells. Here we describe S-Trimer, a native-like trimeric subunit vaccine candidate for COVID-19 based on Trimer-Tag technology. Immunization of S-Trimer with either AS03 (oil-in-water emulsion) or CpG 1018 (TLR9 agonist) plus alum adjuvants induced high-levels of neutralizing antibodies and Th1-biased cellular immune responses in animal models. Moreover, rhesus macaques immunized with adjuvanted S-Trimer were protected from SARS-CoV-2 challenge compared to vehicle controls, based on clinical observations and reduction of viral loads in lungs. Trimer-Tag may be an important new platform technology for scalable production and rapid development of safe and effective subunit vaccines against current and future emerging RNA viruses.

immunology↗

bioRxiv2020

Candidatus (Ca.) phytoplasma asteris subgroups display distinct disease progression dynamics during the carrot growing season

Aster Yellows phytoplasma (AYp; Candidatus (Ca.) Phytoplasma asteris) is an obligate bacterial pathogen that is the causative agent of multiple diseases in herbaceous plants. While this phytoplasma has been examined in depth for its disease characteristics, knowledge about the spatial and temporal dynamics of pathogen spread is lacking. The phytoplasma is found in plants phloem and is vectored by leafhoppers (Cicadellidae: Hemiptera), including the aster leafhopper, Macrosteles quadrilineatus Forbes. The aster leafhopper is a migratory insect pest that overwinters in the southern United States, and historical data suggest these insects migrate from southern overwintering locations to northern latitudes annually, transmitting and driving phytoplasma infection rates as they migrate. A more in-depth understanding of the spatial, temporal and genetic determinants of Aster Yellows disease progress will lead to better integrated pest management strategies for Aster Yellows disease control. Carrot, Daucus carota L., plots were established at two planting densities in central Wisconsin and monitored during the 2018 growing season for Aster Yellows disease progression. Symptomatic carrots were sampled and assayed for the presence of the Aster Yellows phytoplasma. Aster Yellows disease progression was determined to be significantly associated with calendar date, crop density, location within the field, and phytoplasma subgroup.

ecology↗

bioRxiv2020

Eurycomanone regulates lipid metabolism by activating the cAMP/PKA pathway

Eurycoma longifolia Jack (ELJ) contains mainly alkaloids, and quassinoids, which are the main active ingredients. Eurycomanone (EN), one of the most common quassinoids, is said to have beneficial effects on lipid and glucose metabolism. In this study, we investigated the effects of EN on lipolysis by establishing a high-fat animal model in vivo and evaluated its efficacy as a lipolytic and anti-fatty liver agent. Oil red O staining showed morphological changes of 3T3-L1 preadipocytes after EN treatment and confirmed the inhibitory effects of EN on adipocyte differentiation. The mechanism of EN promotes lipolysis in 3T3-L1 cells was analyzed by immunofluorescence, Western blotting, quantitative real-time PCR and siRNA transfection. In C57BL/6J mice fed a high-fat diet, intragastric administration of EN (5 mg/kg and 10 mg/kg) for two weeks, decreased fat droplet mass and size, and reduced fat accumulation in the liver. Furthermore, EN activated PKA and promoted the PKA/hormone sensitive lipase lipolysis signaling pathway, thereby increasing the release of glycerol and free fatty acids from adipocytes. Our findings indicate the potential of EN as a promising alternative pharmacologic agent for the prevention of obesity.

pharmacology and toxicology↗

bioRxiv2020

Laser induced isolation and cultivation of single microbial cells

Single cell isolation and cultivation play an important role in studying physiology, gene expression and functions of microorganisms. Laser Induced Forward Transfer Technique (LIFT) has been applied to isolate single cells but the cell viability after sorting is unclear. We demonstrate that a three-layer LIFT system could be applied to isolate single cells of Gram-negative (E. coli), Gram-positive (Lactobacillus rhamnosus GG, LGG), and eukaryotic microorganisms (Saccharomyces cerevisiae) and the sorted single cells were able to be cultured. The experiment results showed that the average cultivation recovery rate of the ejected single cells were 58% for Saccharomyces cerevisiae, 22% for E. coli, and 74% for Lactobacillus rhamnosus GG (LGG). The identities of the cultured cells from single cell sorting were confirmed by using colony PCR with 16S-rRNA for bacteria and large unit rRNA for yeast and subsequent sequencing. This precise sorting and cultivation technique of live single microbial cells can be coupled with other microscopic approaches (e.g. fluorescent and Raman microscopy) to culture single microorganisms with specific functions, revealing their roles in the natural community. ImportanceSingle cell isolation and cultivation are crucial to recover microorganisms for the study of physiology, gene expression and functions. We developed a laser induced cell sorting technology to precisely isolate single microbial cells from a microscopic slide. More importantly, the isolated single microbial cells are still viable for cultivation. We demonstrate to apply the live sorting method to isolate and cultivate single cells of Gram-negative (E. coli), Gram-positive (Lactobacillus rhamnosus GG, LGG), and eukaryotic microorganisms (Saccharomyces cerevisiae). This precise sorting and cultivation technique can be coupled with other microscopic approaches (e.g. fluorescent and Raman microscopy) to culture specifically targeted single microorganisms from microbial community. Abstract Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/224287v1_ufig1.gif" ALT="Figure 1"> View larger version (42K): org.highwire.dtl.DTLVardef@3ac141org.highwire.dtl.DTLVardef@165c5cforg.highwire.dtl.DTLVardef@9dc0bborg.highwire.dtl.DTLVardef@d86063_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2020

Massive-scale biological activity-based modeling identifies novel antiviral leads against SARS-CoV-2

The recent global pandemic caused by the new coronavirus SARS-CoV-2 presents an urgent need for new therapeutic candidates. While the importance of traditional in silico approaches such as QSAR in such efforts in unquestionable, these models fundamentally rely on structural similarity to infer biological activity and are thus prone to becoming trapped in the very nearby chemical spaces of already known ligands. For novel and unprecedented threats such as COVID-19 much faster and efficient paradigms must be devised to accelerate the identification of new chemical classes for rapid drug development. Here we report the development of a new biological activity-based modeling (BABM) approach that builds on the hypothesis that compounds with similar activity patterns tend to share similar targets or mechanisms of action. In BABM, compound activity profiles established on massive scale across multiple assays are used as signatures to predict compound activity in a new assay or against a new target. We first trained and validated this approach by identifying new antiviral lead candidates for Zika and Ebola based on data from ~0.5 million compounds screened against ~2,000 assays. BABM models were then applied to predict ~300 compounds not previously reported to have activity for SARS-CoV-2, which were then tested in a live virus assay with high (>30%) hit rates. The most potent compounds showed antiviral activities in the nanomolar range. These potent confirmed compounds have the potential to be further developed in novel chemical space into new anti-SARS-CoV-2 therapies. These results demonstrate unprecedented ability using BABM to predict novel structures as chemical leads significantly beyond traditional methods, and its application in rapid drug discovery response in a global public health crisis.

cell biology↗

bioRxiv2020

Platinum resistance induces diverse evolutionary trajectories in high grade serous ovarian cancer

Resistance to therapy is an enduring challenge in cancer care. Here we interrogate this critical unmet need using high grade serous ovarian cancer (HGSC) as a disease model. We have generated a unique panel of platinum-resistant HGSC models and shown that they share multiple transcriptomic features with relapsed human HGSC. Moreover, they evolve diverse in vivo phenotypes reflecting the human disease. We previously characterised copy number signatures in HGSC that correlate with patient survival and now provide the first evidence that these signatures undergo recurrent alterations during platinum therapy. Furthermore, specific, resistance-associated signature change is associated with functionally relevant gene expression differences. For example, reduced signature 3 (BRCA1/2-related homologous recombination deficiency) is associated with increased expression of homologous recombination repair genes (Rad51C, Rad51D, BRCA1) and DNA recombination pathway enrichment. Our mechanistic examination therefore provides new and clinically relevant insights into the genomic evolution of platinum-resistant cancers.

cancer biology↗

bioRxiv2020

Phylogenetic analysis by SNP and development of SSR marker in Passiflora

Information of the Passiflora genome is still very limited. Understand the evolutionary relationship between different species of Passiflora, and develop a large number of SSR markers to provide a basis for the genetic improvement of Passiflora. Applying restriction site associated DNA sequencing (RAD-Seq) technology, we studied the phylogeny, simple sequence repeat (SSR) and marker transferability of 10 accessions of 6 species of Passiflora. Taking the partial assembly sequence of accessions P4 as the reference genome, we constructed the phylogenetic tree using the detected 46,451 high-quality single nucleotide polymorphisms (SNPs), showing that P6, P7, P8 and P9 were a single one while P5 and P10 were clustered together, and P1, P2, P3 and P4 were closer in genetic relationship. Using P8 as the reference genome, a total of 12,452 high-quality SNPs were used to construct phylogenetic tree. P3, P4, P7, P8, P9 and P10 were all single branch while P1 and P2 were clustered together, and P5 and P6 were clustered into one branch. A principal component analysis (PCA) revealed a similar population structure, which four cultivated passion fruits forming a tight cluster. A total of 2,614 SSRs were identified in the genome of 10 Passiflora accessions. The core motifs were AT, GA, AAG etc., 2-6 bases, 4-16 repeats, and 2,515 pairs of SSR primer were successfully developed. Tthe SSR transferability in cultivated passion fruits is the best. These results will contribute to the study of genomics and molecular genetics in passion fruit.

molecular biology↗

bioRxiv2020

Inhibition of ITK differentiates GVT and GVHD in allo-HSCT

Allogeneic hematopoietic stem cell transplantation is a life-saving treatment for many malignant and nonmalignant diseases. Donor T cells contained within the graft prevent tumor recurrence via graft-versus-tumor (GVT) effects, however, also cause graft-versus-host disease (GVHD). Novel treatment strategies are therefore needed to allow maintenance of GVT while suppressing GVHD. Here we show using murine models, that targeting IL-2-inducible T cell kinase (ITK) in donor T cells reduces GVHD while preserving the beneficial GVT effects. Donor T cells from Itk-/- mice exhibit significantly reduced production of inflammatory cytokines and migration to GVHD target organs such as liver and small intestine, while maintaining GVT efficacy against primary B-ALL tumors. Itk-/- T cells exhibited reduced expression of IRF4 and decreased JAK/STAT signaling activity, but preserved cytotoxicity, which was accompanied by upregulation of Eomesodermin (Eomes), which was necessary for GVT function. A novel peptide inhibitor ITK signaling is also able to prevent GVHD. This novel peptide inhibitor also reduced cytokine production in mice and human T cells. Altogether, our data suggest that inhibiting ITK could be a therapeutic strategy to reduce GVHD while preserving the beneficial GVT effects following allo-HSCT treatment. Key PointsO_LIInhibiting ITK by a novel peptide significantly reduces GVHD but retains GVT. C_LIO_LIITK deficient donor T cells exhibit minimal GVHD, but maintain GVT activity. C_LIO_LIITK deficient donor T cells exhibit significantly reduced production of inflammatory cytokines and migration to GVHD target organs. C_LIO_LIEomes is required for GVT effect. C_LI

immunology↗

bioRxiv2020

Targeting heparan sulfate proteoglycan-assisted endocytosis as a COVID-19 therapeutic option

The cell entry of SARS-CoV-2 has emerged as an attractive drug repurposing target for COVID-19. Here we combine genetics and chemical perturbation to demonstrate that ACE2-mediated entry of SARS-CoV and CoV-2 requires the cell surface heparan sulfate (HS) as an assisting cofactor: ablation of genes involved in HS biosynthesis or incubating cells with a HS mimetic both inhibit Spike-mediated viral entry. We show that heparin/HS binds to Spike directly, facilitates the attachment of viral particles to the cell surface to promote cell entry. We screened approved drugs and identified two classes of inhibitors that act via distinct mechanisms to target this entry pathway. Among the drugs characterized, Mitoxantrone is a potent HS inhibitor, while Sunitinib and BNTX disrupt the actin network to indirectly abrogate HS-assisted viral entry. We further show that drugs of the two classes can be combined to generate a synergized activity against SARS-CoV-2-induced cytopathic effect. Altogether, our study establishes HS as an attachment factor that assists SARS coronavirus cell entry, and reveals drugs capable of targeting this important step in the viral life cycle.

cell biology↗

bioRxiv2020

The glycosyltransferase UGT76B1 is critical for plant immunity as it governs the homeostasis of N-hydroxy-pipecolic acid

The trade-off between growth and defense is a critical aspect of plant immunity. Therefore, plant immune response needs to be tightly regulated. The hormone regulating plant defense against biotrophic pathogens is salicylic acid (SA). Recently, N-hydroxy-pipecolic acid (NHP) was identified as second regulator for plant innate immunity and systemic acquired resistance. Although the biosynthetic pathway leading to NHP formation has already been identified, the route how NHP is further metabolized was unclear. Here, we present UGT76B1 as a UDP-dependent glycosyltransferase that modifies NHP by catalyzing the formation of 1-O-glucosyl-pipecolic acid (NHP-OGlc). Analysis of T-DNA and CRISPR knock-out mutant lines of UGT76B1 by targeted and non-targeted UHPLC-HRMS underlined NHP and SA as endogenous substrates of this enzyme in response to Pseudomonas infection and UV treatment. UGT76B1 shows similar KM for NHP and SA. ugt76b1 mutant plants have a dwarf phenotype and a constitutive defense response which can be suppressed by loss of function of the NHP biosynthetic enzyme FMO1. This suggests that elevated accumulation of NHP contributes to the enhanced disease resistance in ugt76b1. Externally applied NHP can move to distal tissue in ugt76b1 mutant plants. Although glycosylation is not required for the long distance movement of NHP during systemic acquired resistance, it is crucial to balance growth and defense.Competing Interest StatementThe authors have declared no competing interest.View Full Text

plant biology↗