Aminopeptidase N is an entry co-factor triggering porcine deltacoronavirus entry via an endocytotic pathway

Porcine deltacoronavirus (PDCoV) is a recently discovered coronavirus that poses a potential threat to the global swine industry. Although we know that aminopeptidase N (APN) is important for PDCoV replication, it is unclear whether it is the primary functional receptor, and the mechanism by which it promotes viral replication is not fully understood. Here, we systematically investigated the role of porcine APN (pAPN) during PDCoV infection of non-susceptible cells, including in viral attachment and internalization. Using a viral entry assay, we found that PDCoV can enter non-susceptible cells but then fails to initiate efficient replication. pAPN and PDCoV virions clearly co-localized with the endocytotic markers RAB5, RAB7, and LAMP1, suggesting that pAPN mediates PDCoV entry by an endocytotic pathway. Most importantly, our study shows that regardless of which receptor PDCoV engages, only entry by an endocytotic route ultimately leads to efficient viral replication. This knowledge should contribute to the development of efficient antiviral treatments, which are especially useful in preventing cross-species transmission. IMPORTANCE PDCoV is a pathogen with potential for transmission across diverse species, although the mechanism of such host-switching events (from swine to other species) is poorly understood. Here, we show that PDCoV enters non-susceptible cells, but without efficient replication. We also investigated the key role played by aminopeptidase N in mediating PDCoV entry via an endocytotic pathway. Our results demonstrate that viral entry via endocytosis is a major determinant of efficient PDCoV replication. This knowledge provides a basis for future studies of the cross-species transmissibility of PDCoV and the development of appropriate anti-viral drugs.

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Meclizine Inhibits Pseudorabies Virus Replication by Interfering With Virus Entry and Release

Frontiers in Microbiology ◽

10.3389/fmicb.2021.795593 ◽

2021 ◽

Vol 12 ◽

Author(s):

Panrao Liu ◽

Danhe Hu ◽

Lili Yuan ◽

Zhengmin Lian ◽

Xiaohui Yao ◽

...

Keyword(s):

Viral Entry ◽

Pseudorabies Virus ◽

Molecular Mechanisms ◽

Clinical Symptoms ◽

Antiviral Effect ◽

Inhibitory Effect ◽

Economic Losses ◽

Swine Industry ◽

Viral Attachment ◽

Viral Loads

Pseudorabies virus (PRV) is a pathogen that causes substantial economic losses to the swine industry. With the emergence and widespread of PRV variants since 2011 in China, current commercial vaccines cannot provide complete protection against PRV infection. Therefore, antiviral drugs may work as an alternative way to control and prevent PRV. In this study, the inhibitory effects and underlying molecular mechanisms of meclizine against PRV were studied. Meclizine displayed a significant inhibitory effect against PRV when it was added before, simultaneously with, or after virus infection. The inhibitory effect of meclizine occurred during viral entry and cell-to-cell spreading but not at viral attachment into PK-15 cells. Meclizine also inhibited viral particle release at the late stage of infection. The antiviral effect of meclizine was tested in mice, and the results showed that meclizine reduced the severity of clinical symptoms and the viral loads in tissues, and delayed the death, after PRV challenge. The above results indicated that meclizine had an inhibitory effect on PRV. Our findings will contribute to the development of potential therapeutic drugs against PRV infection.

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Comprehensive characterization of N- and O- glycosylation of SARS-CoV-2 human receptor angiotensin converting enzyme 2

10.1101/2020.05.01.071688 ◽

2020 ◽

Cited By ~ 12

Author(s):

Asif Shajahan ◽

Stephanie Archer-Hartmann ◽

Nitin T. Supekar ◽

Anne S. Gleinich ◽

Christian Heiss ◽

...

Keyword(s):

Angiotensin Converting Enzyme ◽

Viral Entry ◽

Glycosylation Site ◽

Converting Enzyme ◽

S Protein ◽

Angiotensin Converting Enzyme 2 ◽

Viral Attachment ◽

Viral Binding ◽

Glycosylation Sites

AbstractThe emergence of the COVID-19 pandemic caused by SARS-CoV-2 has created the need for development of new therapeutic strategies. Understanding the mode of viral attachment, entry and replication has become a key aspect of such interventions. The coronavirus surface features a trimeric spike (S) protein that is essential for viral attachment, entry and membrane fusion. The S protein of SARS-CoV-2 binds to human angiotensin converting enzyme 2 (hACE2) for entry. Herein, we describe glycomic and glycoproteomic analysis of hACE2 expressed in HEK293 human cells. We observed high glycan occupancy (73.2 to 100%) at all seven possible N-glycosylation sites and surprisingly detected one novel O-glycosylation site. To deduce the detailed structure of glycan epitopes on hACE2 that may be involved in viral binding, we have characterized the terminal sialic acid linkages, the presence of bisecting GlcNAc, and the pattern of N-glycan fucosylation. We have conducted extensive manual interpretation of each glycopeptide and glycan spectrum, in addition to using bioinformatics tools to validate the hACE2 glycosylation. Our elucidation of the site-specific glycosylation and its terminal orientations on the hACE2 receptor, along with the modeling of hACE2 glycosylation sites can aid in understanding the intriguing virus-receptor interactions and assist in the development of novel therapeutics to prevent viral entry. The relevance of studying the role of ACE2 is further increased due to some recent reports about the varying ACE2 dependent complications with regard to age, sex, race, and pre-existing conditions of COVID-19 patients.

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Polyamines mediate enterovirus attachment directly and indirectly through cellular heparan sulfate synthesis.

10.1101/2021.10.31.466121 ◽

2021 ◽

Author(s):

Bridget M Hulsebosch ◽

Oreoluwa S Omoba ◽

Natalie J LoMascolo ◽

Bryan C. Mounce

Keyword(s):

Heparan Sulfate ◽

Viral Entry ◽

Mammalian Cells ◽

Initiation Factor ◽

Direct Role ◽

Viral Attachment ◽

Cellular Expression ◽

Cellular Translation ◽

Heparan Sulfates

Productive viral infection begins with attachment to a susceptible cell, and viruses have evolved complex mechanisms to attach to and subsequently enter cells. Prior to engagement with a cellular receptor, viruses frequently interact with nonspecific attachment factors that can facilitate virus-receptor interactions and viral entry. Polyamines, small positively-charged molecules abundant in mammalian cells, mediate viral attachment, though the mechanism was not fully understood. Using the Coxsackievirus B3 (CVB3) enterovirus model system, we show that polyamines mediate viral attachment both directly and indirectly. The polyamine putrescine specifically enhances viral attachment to cells depleted of polyamines. Putrescine's positive charge mediates its ability to enhance viral attachment, and polyamine analogs are less efficient at mediating viral attachment. In addition to this direct role of polyamines in attachment, polyamines facilitate the cellular expression of heparan sulfates, negatively-charged molecules found on the cell surface. In polyamine-depleted cells, heparan sulfates are depleted from the surface of cells, resulting in reduced viral attachment. We find that this is due to polyamines' role in the process of hypusination of eukaryotic initiation factor 5A, which facilitates cellular translation. These data highlight the important role of polyamines in mediating cellular attachment, as well as their function in facilitating cellular heparan sulfate synthesis.

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Phytochemical Characterization of Olea europea Leaf Extracts and Assessment of Their Anti-Microbial and Anti-HSV-1 Activity

Viruses ◽

10.3390/v13061085 ◽

2021 ◽

Vol 13 (6) ◽

pp. 1085

Author(s):

Ichrak Ben-Amor ◽

Maria Musarra-Pizzo ◽

Antonella Smeriglio ◽

Manuela D’Arrigo ◽

Rosamaria Pennisi ◽

...

Keyword(s):

Antiviral Activity ◽

Viral Entry ◽

Antiviral Effect ◽

Vero Cells ◽

Biological Properties ◽

Leaf Extracts ◽

Gram Positive Bacteria ◽

Entry Assay ◽

Olea Europea ◽

Hsv 1

Owing to the richness of bioactive compounds, Olea europea leaf extracts exhibit a range of health effects. The present research evaluated the antibacterial and antiviral effect of leaf extracts obtained from Olea europea L. var. sativa (OESA) and Olea europea var. sylvestris (OESY) from Tunisia. LC-DAD-ESI-MS analysis allowed the identification of different compounds that contributed to the observed biological properties. Both OESA and OESY were active against Gram-positive bacteria (MIC values between 7.81 and 15.61 μg/mL and between 15.61 and 31.25 μg/mL against Staphylococcus aureus ATCC 6538 for OESY and OESA, respectively). The antiviral activity against the herpes simplex type 1 (HSV-1) was assessed on Vero cells. The results of cell viability indicated that Olea europea leaf extracts were not toxic to cultured Vero cells. The half maximal cytotoxic concentration (CC50) values for OESA and OESY were 0.2 mg/mL and 0.82 mg/mL, respectively. Furthermore, both a plaque reduction assay and viral entry assay were used to demonstrate the antiviral activity. In conclusion, Olea europea leaf extracts demonstrated a bacteriostatic effect, as well as remarkable antiviral activity, which could provide an alternative treatment against resistant strains.

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Trace Elements as Immunoregulators in SARS-CoV-2 and Other Viral Infections

Indian Journal of Clinical Biochemistry ◽

10.1007/s12291-021-00961-6 ◽

2021 ◽

Author(s):

Karthick Dharmalingam ◽

Amandeep Birdi ◽

Sojit Tomo ◽

Karli Sreenivasulu ◽

Jaykaran Charan ◽

...

Keyword(s):

Immune Response ◽

Trace Elements ◽

Viral Entry ◽

Viral Infections ◽

Inhibitory Effect ◽

Antioxidants Activity ◽

Host Immune Responses ◽

Complex Interactions ◽

Downstream Processes

AbstractNutritional deficiency is associated with impaired immunity and increased susceptibility to infections. The complex interactions of trace elements with the macromolecules trigger the effective immune response against the viral diseases. The outcome of various viral infections along with susceptibility is affected by trace elements such as zinc, selenium, iron, copper, etc. due to their immuno-modulatory effects. Available electronic databases have been comprehensively searched for articles published with full text available and with the key words “Trace elements”, “COVID-19”, “Viral Infections” and “Immune Response” (i.e. separately Zn, Se, Fe, Cu, Mn, Mo, Cr, Li, Ni, Co) appearing in the title and abstract. On the basis of available articles we have explored the role of trace elements in viral infections with special reference to COVID-19 and their interactions with the immune system. Zinc, selenium and other trace elements are vital to triggerTH1 cells and cytokine-mediated immune response for substantial production of proinflammatory cytokines. The antiviral activity of some trace elements is attributed to their inhibitory effect on viral entry, replication and other downstream processes. Trace elements having antioxidants activity not only regulate host immune responses, but also modify the viral genome. Adequate dietary intake of trace elements is essential for activation, development, differentiation and numerous functions.

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Chloroplast Hsp70 Isoform Is Required for Age-Dependent Tissue Preference of Bamboo mosaic virus in Mature Nicotiana benthamiana Leaves

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-01-17-0012-r ◽

2017 ◽

Vol 30 (8) ◽

pp. 631-645 ◽

Cited By ~ 3

Author(s):

Ying Wen Huang ◽

Chung Chi Hu ◽

Ching Hsiu Tsai ◽

Na Sheng Lin ◽

Yau Heiu Hsu

Keyword(s):

Mosaic Virus ◽

Nicotiana Benthamiana ◽

Transit Peptide ◽

Plant Viruses ◽

Age Dependent ◽

Efficient Replication ◽

Underlying Mechanisms ◽

Lines Of Evidence ◽

Suitable Environment

Plant viruses may exhibit age-dependent tissue preference in their hosts but the underlying mechanisms are not well understood. In this study, we provide several lines of evidence to reveal the determining role of a protein of the Nicotiana benthamiana chloroplast Hsp70 (NbcpHsp70) family, NbcpHsp70-2, involved in the preference of Bamboo mosaic virus (BaMV) to infect older tissues. NbcpHsp70 family proteins were identified in complexes pulled down with BaMV replicase as the bait. Among the isoforms of NbcpHsp70, only the specific silencing of NbcpHsp70-2 resulted in the significant decrease of BaMV RNA in N. benthamiana protopalsts, indicating that NbcpHsp70-2 is involved in the efficient replication of BaMV RNA. We further identified the age-dependent import regulation signal contained in the transit peptide of NbcpHsp70-2. Deletion, overexpression, and substitution experiments revealed that the signal in the transit peptide of NbcpHsp70-2 is crucial for both the import of NbcpHsp70-2 into older chloroplasts and the preference of BaMV for infecting older leaves of N. benthamiana. Together, these data demonstrated that BaMV may exploit a cellular age-dependent transportation mechanism to target a suitable environment for viral replication.

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Synergistic In Vitro Antiretroviral Activity of a Humanized Monoclonal Anti-CD4 Antibody (TNX-355) and Enfuvirtide (T-20)

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00761-05 ◽

2006 ◽

Vol 50 (6) ◽

pp. 2231-2233 ◽

Cited By ~ 39

Author(s):

Xing-Quan Zhang ◽

Meredith Sorensen ◽

Michael Fung ◽

Robert T. Schooley

Keyword(s):

Viral Replication ◽

Viral Entry ◽

Antiretroviral Agents ◽

Entry Inhibitors ◽

High Level ◽

Antiretroviral Activity

ABSTRACT Recently, antiretroviral agents directed at several steps involved in viral entry have been shown to reduce viral replication in vitro and in vivo. We have demonstrated a high level of in vitro synergistic antiretroviral activity for two entry inhibitors that are directed at sequential steps in the entry process.

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HIV-1 requires Arf6-mediated membrane dynamics to efficiently enter and infect T lymphocytes

Molecular Biology of the Cell ◽

10.1091/mbc.e10-08-0722 ◽

2011 ◽

Vol 22 (8) ◽

pp. 1148-1166 ◽

Cited By ~ 22

Author(s):

Laura García-Expósito ◽

Jonathan Barroso-González ◽

Isabel Puigdomènech ◽

José-David Machado ◽

Julià Blanco ◽

...

Keyword(s):

Plasma Membrane ◽

T Lymphocytes ◽

Viral Entry ◽

Membrane Trafficking ◽

Membrane Dynamics ◽

Viral Fusion ◽

Viral Attachment ◽

Immunodeficiency Virus ◽

Vesicular Stomatitis ◽

Hiv 1

As the initial barrier to viral entry, the plasma membrane along with the membrane trafficking machinery and cytoskeleton are of fundamental importance in the viral cycle. However, little is known about the contribution of plasma membrane dynamics during early human immunodeficiency virus type 1 (HIV-1) infection. Considering that ADP ribosylation factor 6 (Arf6) regulates cellular invasion via several microorganisms by coordinating membrane trafficking, our aim was to study the function of Arf6-mediated membrane dynamics on HIV-1 entry and infection of T lymphocytes. We observed that an alteration of the Arf6–guanosine 5′-diphosphate/guanosine 5′-triphosphate (GTP/GDP) cycle, by GDP-bound or GTP-bound inactive mutants or by specific Arf6 silencing, inhibited HIV-1 envelope–induced membrane fusion, entry, and infection of T lymphocytes and permissive cells, regardless of viral tropism. Furthermore, cell-to-cell HIV-1 transmission of primary human CD4+T lymphocytes was inhibited by Arf6 knockdown. Total internal reflection fluorescence microscopy showed that Arf6 mutants provoked the accumulation of phosphatidylinositol-(4,5)-biphosphate–associated structures on the plasma membrane of permissive cells, without affecting CD4-viral attachment but impeding CD4-dependent HIV-1 entry. Arf6 silencing or its mutants did not affect fusion, entry, and infection of vesicular stomatitis virus G–pseudotyped viruses or ligand-induced CXCR4 or CCR5 endocytosis, both clathrin-dependent processes. Therefore we propose that efficient early HIV-1 infection of CD4+T lymphocytes requires Arf6-coordinated plasma membrane dynamics that promote viral fusion and entry.

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