scholarly journals Imaging Viral Infection by Fluorescence Microscopy: Focus on HIV-1 Early Stage

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 213
Author(s):  
Soumajit Mukherjee ◽  
Emmanuel Boutant ◽  
Eleonore Réal ◽  
Yves Mély ◽  
Halina Anton
Keyword(s):  
Host Cell ◽  
Viral Entry ◽  
Integration Site ◽  
Viral Envelope ◽  
Integration Step ◽  
Imaging Studies ◽  
Cytoplasmic Transport ◽  
Capsid Shell ◽  
Wide Range ◽  
Hiv 1

During the last two decades, progresses in bioimaging and the development of various strategies to fluorescently label the viral components opened a wide range of possibilities to visualize the early phase of Human Immunodeficiency Virus 1 (HIV-1) life cycle directly in infected cells. After fusion of the viral envelope with the cell membrane, the viral core is released into the cytoplasm and the viral RNA (vRNA) is retro-transcribed into DNA by the reverse transcriptase. During this process, the RNA-based viral complex transforms into a pre-integration complex (PIC), composed of the viral genomic DNA (vDNA) coated with viral and host cellular proteins. The protective capsid shell disassembles during a process called uncoating. The viral genome is transported into the cell nucleus and integrates into the host cell chromatin. Unlike biochemical approaches that provide global data about the whole population of viral particles, imaging techniques enable following individual viruses on a single particle level. In this context, quantitative microscopy has brought original data shedding light on the dynamics of the viral entry into the host cell, the cytoplasmic transport, the nuclear import, and the selection of the integration site. In parallel, multi-color imaging studies have elucidated the mechanism of action of host cell factors implicated in HIV-1 viral cycle progression. In this review, we describe the labeling strategies used for HIV-1 fluorescence imaging and report on the main advancements that imaging studies have brought in the understanding of the infection mechanisms from the viral entry into the host cell until the provirus integration step.

Recent Progress in the Development of HIV-1 Entry Inhibitors: From Small Molecules to Potent Anti-HIV Agents

2019 ◽  
Vol 19 (18) ◽  
pp. 1599-1620 ◽  
Author(s):  
Khomson Suttisintong ◽  
Narongpol Kaewchangwat ◽  
Eknarin Thanayupong ◽  
Chakkrapan Nerungsi ◽  
Onsiri Srikun ◽  
...  
Keyword(s):  
Host Cell ◽  
Viral Entry ◽  
Chemokine Receptors ◽  
Recent Progress ◽  
Viral Envelope ◽  
Surface Glycoprotein ◽  
Entry Inhibitors ◽  
Transmembrane Glycoprotein ◽  
Glycoprotein Gp120 ◽  
Hiv 1

Viral entry, the first process in the reproduction of viruses, primarily involves attachment of the viral envelope proteins to membranes of the host cell. The crucial components that play an important role in viral entry include viral surface glycoprotein gp120, viral transmembrane glycoprotein gp41, host cell glycoprotein (CD4), and host cell chemokine receptors (CCR5 and CXCR4). Inhibition of the multiple molecular interactions of these components can restrain viruses, such as HIV-1, from fusion with the host cell, blocking them from reproducing. This review article specifically focuses on the recent progress in the development of small-molecule HIV-1 entry inhibitors and incorporates important aspects of their structural modification that lead to the discovery of new molecular scaffolds with more potency.

scholarly journals HIV Cell Fusion Assay

2013 ◽  
Vol 19 (1) ◽  
pp. 108-118 ◽  
Author(s):  
Elizabeth B. Smith ◽  
Robert A. Ogert ◽  
David Pechter ◽  
Artjohn Villafania ◽  
Susan J. Abbondanzo ◽  
...  
Keyword(s):  
Host Cell ◽  
Cell Fusion ◽  
Chemokine Receptor ◽  
Viral Entry ◽  
Envelope Glycoprotein ◽  
Laser Scanning ◽  
Viral Envelope ◽  
Fusion Event ◽  
Cxcr4 Signaling ◽  
Hiv Entry

The health and disease-related biology of the CXCR4 chemokine receptor presents the challenge of finding a small molecule that can bind CXCR4 and block T-cell tropic human immunodeficiency virus type 1 (HIV-1) cell entry, while preserving the ability of CXCR4 to respond to its native ligand, CXCL12. HIV entry into the host cell involves the interaction of the viral envelope glycoprotein gp120 binding to CD4, followed by a rearrangement in gp120, and subsequent interaction with the chemokine receptor CXCR4 or CCR5. These initial events can be re-created in a cell fusion assay that represents a surrogate system, mimicking the early stages of viral entry via these host cell receptors. In the current study, a T-tropic HIV cell fusion assay was established using U2OS cells expressing the envelope glycoprotein gp160 from the T-tropic HIV NL4-3 and HeLa cells expressing CD4 and CXCR4. Detection of the cell fusion event was based on a Gal4/VP16-activated β-lactamase signal and was measured by automated microscopy or laser scanning plate cytometry. Changes in morphology associated with cell fusion were combined with β-lactamase activity to generate results with robust assay statistics in both 384-well and 1536-well plates. Compounds were subsequently characterized by CXCR4 signaling assays to eliminate functional antagonists and allow the identification of a function-sparing HIV entry inhibitor.

scholarly journals Development of a generative adversarial neural network for identification of potential HIV-1 inhibitors by deep learning methods

Informatics ◽  
2020 ◽  
Vol 17 (1) ◽  
pp. 7-17
Author(s):  
G. I. Nikolaev ◽  
N. A. Shuldov ◽  
A. I. Anishenko, ◽  
A. V. Tuzikov ◽  
A. M. Andrianov
Keyword(s):  
Neural Network ◽  
Deep Learning ◽  
Rational Design ◽  
Binding Free Energy ◽  
Viral Envelope ◽  
Training Dataset ◽  
Learning Methods ◽  
The Neural Network ◽  
Wide Range ◽  
Hiv 1

A generative adversarial autoencoder for the rational design of potential HIV-1 entry inhibitors able to block the region of the viral envelope protein gp120 critical for the virus binding to cellular receptor CD4 was developed using deep learning methods. The research were carried out to create the  architecture of the neural network, to form  virtual compound library of potential anti-HIV-1 agents for training the neural network, to make  molecular docking of all compounds from this library with gp120, to  calculate the values of binding free energy, to generate molecular fingerprints for chemical compounds from the training dataset. The training the neural network was implemented followed by estimation of the learning outcomes and work of the autoencoder.  The validation of the neural network on a wide range of compounds from the ZINC database was carried out. The use of the neural network in combination with virtual screening of chemical databases was shown to form a productive platform for identifying the basic structures promising for the design of novel antiviral drugs that inhibit the early stages of HIV infection.

scholarly journals HIV-1 Envelope Glycoprotein at the Interface of Host Restriction and Virus Evasion

Viruses ◽  
2019 ◽  
Vol 11 (4) ◽  
pp. 311 ◽  
Author(s):  
Saina Beitari ◽  
Yimeng Wang ◽  
Shan-Lu Liu ◽  
Chen Liang
Keyword(s):  
Viral Infection ◽  
Adaptive Immunity ◽  
Viral Entry ◽  
Envelope Protein ◽  
Envelope Proteins ◽  
Viral Envelope ◽  
Innate Immune ◽  
Host Restriction ◽  
Hiv 1 ◽  
Viral Envelope Proteins

Without viral envelope proteins, viruses cannot enter cells to start infection. As the major viral proteins present on the surface of virions, viral envelope proteins are a prominent target of the host immune system in preventing and ultimately eliminating viral infection. In addition to the well-appreciated adaptive immunity that produces envelope protein-specific antibodies and T cell responses, recent studies have begun to unveil a rich layer of host innate immune mechanisms restricting viral entry. This review focuses on the exciting progress that has been made in this new direction of research, by discussing various known examples of host restriction of viral entry, and diverse viral countering strategies, in particular, the emerging role of viral envelope proteins in evading host innate immune suppression. We will also highlight the effective cooperation between innate and adaptive immunity to achieve the synergistic control of viral infection by targeting viral envelope protein and checking viral escape. Given that many of the related findings were made with HIV-1, we will use HIV-1 as the model virus to illustrate the basic principles and molecular mechanisms on host restriction targeting HIV-1 envelope protein.

scholarly journals Low pH Is Required for Avian Sarcoma and Leukosis Virus Env-Dependent Viral Penetration into the Cytosol and Not for Viral Uncoating

2004 ◽  
Vol 78 (19) ◽  
pp. 10433-10441 ◽  
Author(s):  
Richard J. O. Barnard ◽  
Shakti Narayan ◽  
Geethanjali Dornadula ◽  
Michael D. Miller ◽  
John A. T. Young
Keyword(s):  
Viral Entry ◽  
Strong Support ◽  
Low Ph ◽  
Viral Envelope ◽  
Specific Cell ◽  
Dependent Manner ◽  
Surface Receptors ◽  
Ph Dependent ◽  
Avian Sarcoma ◽  
Hiv 1

ABSTRACT A novel entry mechanism has been proposed for the avian sarcoma and leukosis virus (ASLV), whereby interaction with specific cell surface receptors activates or primes the viral envelope glycoprotein (Env), rendering it sensitive to subsequent low-pH-dependent fusion triggering in acidic intracellular organelles. However, ASLV fusion seems to proceed to a lipid mixing stage at neutral pH, leading to the suggestion that low pH might instead be required for a later stage of viral entry such as uncoating (L. J. Earp, S. E. Delos, R. C. Netter, P. Bates, and J. M. White. J. Virol. 77:3058-3066, 2003). To address this possibility, hybrid virus particles were generated with the core of human immunodeficiency virus type 1 (HIV-1), a known pH-independent virus, and with subgroups A or B ASLV Env proteins. Infection of cells by these pseudotyped virions was blocked by lysosomotropic agents, as judged by inhibition of HIV-1 DNA synthesis. Furthermore, by using HIV-1 cores that contain a Vpr-β-lactamase fusion protein (Vpr-BlaM) to monitor viral penetration into the cytosol, we demonstrated that virions bearing ASLV Env, but not HIV-1 Env, enter the cytosol in a low-pH-dependent manner. This effect was independent of the presence of the cytoplasmic tail of ASLV Env. These studies provide strong support for the model, indicating that low pH is required for ASLV Env-dependent viral penetration into the cytosol and not for viral uncoating.

scholarly journals gp120 Alters Its Conformation to Enhance Evasiveness and Infectivity

2021 ◽  
Author(s):  
Joseph A. Ayariga ◽  
Logan Gildea ◽  
Ayodeji Ipimoroti ◽  
Qiana L. Matthews
Keyword(s):  
Structural Dynamics ◽  
Viral Entry ◽  
Antiviral Agents ◽  
Host Immunity ◽  
Viral Envelope ◽  
Cell Surface Receptor ◽  
Type I ◽  
Structural Constraints ◽  
Conformational States ◽  
Hiv 1

Infection by human immunodeficiency virus type I (HIV-1) requires virus particle binding to host cell-surface receptor CD4 via the viral envelope glycoprotein gp120. HIV-1 therapy and prevention efforts involve development of mimetic or recombinant gp120 vaccines or deployment of antiviral agents that target specific epitopes of gp120. The unliganded conformational state of gp120 is closed, whereas the CD4-bound state is open. However, in between, there exist dynamic conformational states, indicating intrinsically flexible region(s) of structural dynamics, imposing a structural challenge for developing drug or antibody targets. Known conformational states of gp120 were determined by X-ray crystallographic and cryo-electron microscopy, and neither method captures the population of gp120 species arising from conformational plasticity, motions, and transitions. gp120 plasticity brings up several important questions. How will differences in conformation affect receptor binding, antibody recognition, and neutralization? Which regions are crucial for gp120 structural plasticity? How could structural dynamics influence HIV-1 evasiveness against host immunity and drugs or vaccines, and facilitate the viral entry into its host? This review explores the structural constraints presented by conformational states of the glycoprotein to antibodies or drugs and how these conformational states provide structural avenues for the virus to escape neutralizing agents and evade host immunity.

scholarly journals Antibody Binding Is a Dominant Determinant of the Efficiency of Human Immunodeficiency Virus Type 1 Neutralization

2006 ◽  
Vol 80 (22) ◽  
pp. 11404-11408 ◽  
Author(s):  
Xinzhen Yang ◽  
Inna Lipchina ◽  
Simon Cocklin ◽  
Irwin Chaiken ◽  
Joseph Sodroski
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Antibody Binding ◽  
Viral Envelope ◽  
Envelope Glycoproteins ◽  
Wide Range ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Primary and laboratory-adapted variants of human immunodeficiency virus type 1 (HIV-1) exhibit a wide range of sensitivities to neutralization by antibodies directed against the viral envelope glycoproteins. An antibody directed against an artificial FLAG epitope inserted into the envelope glycoproteins of three HIV-1 isolates with vastly different neutralization sensitivities inhibited all three viruses equivalently. Thus, naturally occurring HIV-1 isolates that are neutralization resistant are not necessarily more impervious to the inhibitory consequences of bound antibody. Moreover, the binding affinity of the anti-FLAG antibody correlated with neutralizing potency, underscoring the dominant impact on neutralization of antibody binding to the envelope glycoproteins.

scholarly journals Inhibition of Endosomal/Lysosomal Degradation Increases the Infectivity of Human Immunodeficiency Virus

2002 ◽  
Vol 76 (22) ◽  
pp. 11440-11446 ◽  
Author(s):  
Brenda L. Fredericksen ◽  
Bangdong L. Wei ◽  
Jian Yao ◽  
Tianci Luo ◽  
J. Victor Garcia
Keyword(s):  
Human Immunodeficiency Virus ◽  
Host Cell ◽  
Leukemia Virus ◽  
Fold Increase ◽  
Viral Envelope ◽  
Endocytic Pathway ◽  
Target Cells ◽  
Concanamycin A ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Productive entry of human immunodeficiency virus type 1 (HIV-1) into a host cell is believed to proceed via fusion of the viral envelope with the host cell's plasma membrane. Interestingly, the majority of HIV-1 particles that bind to the cell surface are taken up by the host cell via endocytosis; however, this mode of internalization generally does not result in infection. Presumably, virus particles remain trapped in the endocytic pathway and are eventually degraded. Here, we demonstrate that treatment of cells with various pharmacological agents known to elevate the pH of endosomes and lysosomes allows HIV-1 to efficiently enter and infect the host cell. Pretreatment of cells with bafilomycin A1 results in up to a 50-fold increase in the infectivity of HIV-1SF2. Similarly, pretreatment of target cells with amantadine, concanamycin A, concanamycin B, chloroquine, and ammonium chloride resulted in increases in HIV-1 infectivity ranging between 2- and 15-fold. Analysis of receptor and coreceptor expression, HIV-long terminal repeat (LTR) transactivation, and transduction with amphotropic-pseudotyped murine leukemia virus (MLV)-based vectors suggests that the increase in infectivity is not artifactual. The increased infectivity under these conditions appears to be due to the ability of HIV-1 and MLV particles to enter via the endocytic pathway when spared from degradation in the late endosomes and lysosomes. These results could have significant implications for the administration of current and future lysosmotropic agents to patients with HIV disease.

scholarly journals Fusion Promotion by a Paramyxovirus Hemagglutinin-Neuraminidase Protein: pH Modulation of Receptor Avidity of Binding Sites I and II

2007 ◽  
Vol 81 (17) ◽  
pp. 9152-9161 ◽  
Author(s):  
Laura M. Palermo ◽  
Matteo Porotto ◽  
Olga Greengard ◽  
Anne Moscona
Keyword(s):  
Host Cell ◽  
Receptor Binding ◽  
Viral Entry ◽  
Ph Dependence ◽  
Initial Step ◽  
Viral Envelope ◽  
Respiratory Pathogen ◽  
Receptor Binding Site ◽  
Host Cell Membrane ◽  
Neutral Ph

ABSTRACT Paramyxoviruses, including the childhood respiratory pathogen human parainfluenza virus type 3 (HPIV3), possess an envelope protein hemagglutinin-neuraminidase (HN) that has receptor-cleaving (neuraminidase), as well as receptor-binding, activity. HN is a type II transmembrane glycoprotein, present on the surface of the virus as a tetramer composed of two dimers. HN is also essential for activating the fusion protein (F) to mediate merger of the viral envelope with the host cell membrane. This initial step of viral entry occurs at the host cell surface at neutral pH. The HN molecule carries out these three different critical activities at specific points in the process of viral entry, and understanding the regulation of these activities is key for the design of strategies that block infection. One bifunctional site (site I) on the HN of HPIV3 possesses both receptor binding and neuraminidase activities, and we recently obtained experimental evidence for a second receptor binding site (site II) on HPIV3 HN. Mutation of HN at specific residues at this site, which is next to the HN dimer interface, confers enhanced fusion properties, without affecting neuraminidase activity or receptor binding at neutral pH. We now demonstrate that mutations at this site II, as well as at site I, confer pH dependence on HN′s receptor avidity. These mutations permit pH to modulate the binding and fusion processes of the virus, potentially providing regulation at specific stages of the viral life cycle.

scholarly journals An Envelope-Determined, pH-Independent Endocytic Route of Viral Entry Determines the Susceptibility of Human Immunodeficiency Virus Type 1 (HIV-1) and HIV-2 to Lv2 Restriction

2005 ◽  
Vol 79 (15) ◽  
pp. 9410-9418 ◽  
Author(s):  
David Marchant ◽  
Stuart J. D. Neil ◽  
Keith Aubin ◽  
Christian Schmitz ◽  
Áine McKnight
Keyword(s):  
Human Immunodeficiency Virus ◽  
Lipid Rafts ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Viral Entry ◽  
Viral Envelope ◽  
Endocytic Pathway ◽  
Immunodeficiency Virus ◽  
Cellular Tropism ◽  
Hiv 1

ABSTRACT We identified a postentry restriction, termed Lv2, which determines the cellular tropism of two related human immunodeficiency virus type 2 (HIV-2) isolates and is dependent on the sequence of the capsid (CA) and envelope (Env) proteins. To explain the reliance on both CA and Env, we proposed that restrictive Envs deliver susceptible capsids to a compartment where Lv2 is active whereas nonrestrictive Envs deliver capsids into a compartment where Lv2 is either absent or less active. To test this model, we used compounds that affect endocytic pathways (ammonium chloride, bafilomycin A1, hypertonic sucrose) or lipid rafts (methyl-β-cyclodextrin) to treat restrictive cells and show that restricted virus can be rescued from Lv2 if a lipid-raft-dependent, pH-independent endocytic pathway is inhibited. Furthermore, viral entry into HeLa/CD4 cells containing a tailless CD4 receptor, located outside lipid rafts, was fully permissive. Finally, we show that a variety of primary HIV-1 and HIV-2 viruses are susceptible to Lv2. Thus, we show that the route of entry, determined by the viral envelope, can influence cellular tropism by avoiding intracellular blocks to infection.

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