scholarly journals Caveolae Are Involved in the Trafficking of Mouse Polyomavirus Virions and Artificial VP1 Pseudocapsids toward Cell Nuclei

2001 ◽  
Vol 75 (22) ◽  
pp. 10880-10891 ◽  
Author(s):  
Zuzana Richterová ◽  
David Liebl ◽  
Martin Horák ◽  
Zdena Palková ◽  
Jitka Štokrová ◽  
...  
Keyword(s):  
Viral Particle ◽  
Stress Fibers ◽  
Particle Movement ◽  
Cell Nuclei ◽  
Membrane Structures ◽  
Caveolin 1 ◽  
Strong Signal ◽  
Artificial Virus ◽  
Viral Particles ◽  
Perinuclear Area

ABSTRACT Electron and confocal microscopy were used to observe the entry and the movement of polyomavirus virions and artificial virus-like particles (VP1 pseudocapsids) in mouse fibroblasts and epithelial cells. No visible differences in adsorption and internalization of virions and VP1 pseudocapsids (“empty” or containing DNA) were observed. Viral particles entered cells internalized in smooth monopinocytic vesicles, often in the proximity of larger, caveola-like invaginations. Both “empty” vesicles derived from caveolae and vesicles containing viral particles were stained with the anti-caveolin-1 antibody, and the two types of vesicles often fused in the cytoplasm. Colocalization of VP1 with caveolin-1 was observed during viral particle movement from the plasma membrane throughout the cytoplasm to the perinuclear area. Empty vesicles and vesicles with viral particles moved predominantly along microfilaments. Particle movement was accompanied by transient disorganization of actin stress fibers. Microfilaments decorated by the VP1 immunofluorescent signal could be seen as concentric curves, apparently along membrane structures that probably represent endoplasmic reticulum. Colocalization of VP1 with tubulin was mostly observed in areas close to the cell nuclei and on mitotic tubulin structures. By 3 h postinfection, a strong signal of the VP1 (but no viral particles) had accumulated in the proximity of nuclei, around the outer nuclear membrane. However, the vast majority of VP1 pseudocapsids did not enter the nuclei.

scholarly journals Contribution of the Cytoplasmic Determinants of Vpu to the Expansion of Virus-Containing Compartments in HIV-1-Infected Macrophages

2019 ◽  
Vol 93 (11) ◽  
Author(s):  
Olivier Leymarie ◽  
Leslie Lepont ◽  
Margaux Versapuech ◽  
Delphine Judith ◽  
Sophie Abelanet ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Viral Particle ◽  
Transmembrane Domain ◽  
Immune Surveillance ◽  
Restriction Factor ◽  
Viral Particles ◽  
Protein Functions ◽  
Specific Accumulation ◽  
Model Cell ◽  
Hiv 1

ABSTRACTHIV-1 infection of macrophages leads to the sequestration of newly formed viruses in intracellular plasma membrane-connected structures termed virus-containing compartments (VCCs), where virions remain infectious and hidden from immune surveillance. The cellular restriction factor bone marrow stromal cell antigen 2 (BST2), which prevents HIV-1 dissemination by tethering budding viral particles at the plasma membrane, can be found in VCCs. The HIV-1 accessory protein Vpu counteracts the restriction factor BST2 by downregulating its expression and removing it from viral budding sites. Numerous studies described these Vpu countermeasures in CD4+T cells or model cell lines, but the interplay between Vpu and BST2 in VCC formation and HIV-1 production in macrophages is less explored. Here, we show that Vpu expression in HIV-1-infected macrophages enhances viral release. This effect is related to Vpu’s ability to circumvent BST2 antiviral activity. We show that in absence of Vpu, BST2 is enriched in VCCs and colocalizes with capsid p24, whereas Vpu expression significantly reduces the presence of BST2 in these compartments. Furthermore, our data reveal that BST2 is dispensable for the formation of VCCs and that Vpu expression impacts the volume of these compartments. This Vpu activity partly depends on BST2 expression and requires the integrity of the Vpu transmembrane domain, the dileucine-like motif E59XXXLV64and phosphoserines 52 and 56 of Vpu. Altogether, these results highlight that Vpu controls the volume of VCCs and promotes HIV-1 release from infected macrophages.IMPORTANCEHIV-1 infection of macrophages leads to the sequestration of newly formed viruses in virus-containing compartments (VCCs), where virions remain infectious and hidden from immune surveillance. The restriction factor BST2, which prevents HIV-1 dissemination by tethering budding viral particles, can be found in VCCs. The HIV-1 Vpu protein counteracts BST2. This study explores the interplay between Vpu and BST2 in the viral protein functions on HIV-1 release and viral particle sequestration in VCCs in macrophages. The results show that Vpu controls the volume of VCCs and favors viral particle release. These Vpu functions partly depend on Vpu’s ability to antagonize BST2. This study highlights that the transmembrane domain of Vpu and two motifs of the Vpu cytoplasmic domain are required for these functions. These motifs were notably involved in the control of the volume of VCCs by Vpu but were dispensable for the prevention of the specific accumulation of BST2 in these structures.

scholarly journals Colorimetric Test for Fast Detection of SARS-CoV-2 in Nasal and Throat Swabs

2020 ◽  
Author(s):  
Bartolomeo Della Ventura ◽  
Michele Cennamo ◽  
Antonio Minopoli ◽  
Raffaele Campanile ◽  
Sergio Bolletti Censi ◽  
...  
Keyword(s):  
Viral Particle ◽  
Extinction Spectrum ◽  
Colorimetric Method ◽  
Surface Proteins ◽  
Mixed Solution ◽  
Colorimetric Test ◽  
The People ◽  
Viral Particles ◽  
New Perspective ◽  
Viral Target

Mass testing is fundamental to face the pandemic caused by the coronavirus SARS-CoV-2 discovered at the end of 2019. To this aim, it is necessary to establish reliable, fast and cheap tools to detect viral particles in biological material so to identify the people capable to spread the infection. We demonstrate that a colorimetric biosensor based on gold nanoparticle (AuNP) interaction induced by SARS-CoV-2 lends itself as an outstanding tool for detecting viral particles in nasal and throat swabs. The extinction spectrum of a colloidal solution of multiple viral-target gold nanoparticles - AuNPs functionalized with antibodies targeting three surface proteins of SARS-CoV-2 (spike, envelope and membrane) - is redshifted in few minutes when mixed to a solution containing the viral particle. The optical density of the mixed solution measured at 560 nm was compared to the threshold cycle (Ct) of a Real Time-PCR (gold standard for detecting the presence of viruses) finding that the colorimetric method is able to detect very low viral load with a detection limit approaching that of RT-PCR. Since the method is sensitive to the infecting viral particle rather than to its RNA, the achievements reported here open new perspective not only in the context of the current and possible future pandemics, but also in microbiology as the biosensor proves itself to be a powerful though simple tool for measuring the viral particle concentration.

scholarly journals Functional Correlation between Subcellular Localizations of Japanese Encephalitis Virus Capsid Protein and Virus Production

2019 ◽  
Vol 93 (19) ◽  
Author(s):  
Kotaro Ishida ◽  
Simon Goto ◽  
Marina Ishimura ◽  
Misato Amanuma ◽  
Yumiko Hara ◽  
...  
Keyword(s):  
Japanese Encephalitis Virus ◽  
Virus Particle ◽  
Capsid Protein ◽  
Viral Particle ◽  
Particle Formation ◽  
Molecular Surface ◽  
Functional Correlation ◽  
Viral Particles ◽  
Detergent Treatment ◽  
Virus Capsid Protein

ABSTRACT The flavivirus capsid protein is considered to be essential for the formation of nucleocapsid complexes with viral genomic RNA at the viral replication organelle that appears on the endoplasmic reticulum (ER), as well as for incorporation into virus particles. However, this protein is also detected at the lipid droplet (LD) and nucleolus, and physiological roles of these off-site localizations are still unclear. In this study, we made a series of alanine substitution mutants of Japanese encephalitis virus (JEV) capsid protein that cover all polar and hydrophobic amino acid residues to identify the molecular surfaces required for virus particle formation and for localization at the LD and nucleolus. Five mutants exhibited a defect in the formation of infectious particles, and two of these mutants failed to be incorporated into the subviral particles (SVP). Three mutants lost the ability to localize to the nucleolus, and only a single mutant, with mutations at α2, was unable to localize to the LD. Unlike the cytoplasmic capsid protein, the nucleolar capsid protein was resistant to detergent treatment, and the α2 mutant was hypersensitive to detergent treatment. To scrutinize the relationship between these localizations and viral particle formation, we made eight additional alanine substitution mutants and found that all the mutants that did not localize at the LD or nucleolus failed to form normal viral particles. These results support the functional correlation between LD or nucleolus localization of the flaviviral capsid protein and the formation of infectious viral particles. IMPORTANCE This study is the first to report the comprehensive mutagenesis of a flavivirus capsid protein. We assessed the requirement of each molecular surface for infectious viral particle formation as well as for LD and nucleolar localization and found functional relationships between the subcellular localization of the virus capsid protein and infectious virus particle formation. We developed a system to independently assess the packaging of viral RNA and that of the capsid protein and found a molecular surface of the capsid protein that is crucial for packaging of viral RNA but not for packaging of the capsid protein itself. We also characterized the biochemical properties of capsid protein mutants and found that the capsid protein localizes at the nucleolus in a different manner than for its localization to the LD. Our comprehensive alanine-scanning mutagenesis study will aid in the development of antiflavivirus small molecules that can target the flavivirus capsid protein.

Virus Particles Infected the Erythrocytes of Cultured Eel in Taiwan

1981 ◽  
Vol 39 ◽  
pp. 608-609
Author(s):  
Kuo-chun Liu
Keyword(s):  
Viral Particle ◽  
Electron Microscopic Observation ◽  
Field Trips ◽  
Fish Farms ◽  
Electron Microscopic ◽  
Internal Organs ◽  
Virus Particles ◽  
Pathological Conditions ◽  
Viral Particles ◽  
Electron Lucent

The erythrocytes infected by viral particles and the pathological conditions by the presence of those viral particles were reported in cod, herring and few other species of fish in the western waters. This report will present the evidence that the erythrocytes of some of the cultured eel in Taiwan were infected by virus particles.In the summer of 1980, in one of the field trips to the fish farms in Lotung county, northeastern of Taiwan, the author was informed that dead eels from unknown cause were picked up from eel ponds every morning. Samples were collected the following morning. There was no observable external symptom.The internal organs showed a red coloration (Fig. 1,2). Tissues from the livers and gills were processed for electron microscopic observation. Virus particles were found in some of the erythrocytes in both organs (Fig.3).The viral particles aggregated together as an inclusion body in the cytoplasm only. No intranuclear viral particle was found. In a single viral aggregate 1 to 40 particles were observed. A electron lucent ring separated the viral aggregate from the hemoglobin cytoplasm.

scholarly journals The Tetraspan Protein EMP2 Modulates the Surface Expression of Caveolins and Glycosylphosphatidyl Inositol-linked Proteins

2004 ◽  
Vol 15 (5) ◽  
pp. 2073-2083 ◽  
Author(s):  
Madhuri Wadehra ◽  
Lee Goodglick ◽  
Jonathan Braun
Keyword(s):  
Lipid Rafts ◽  
Surface Expression ◽  
Bacterial Toxin ◽  
Membrane Structures ◽  
Nih3t3 Cells ◽  
Caveolin 1 ◽  
Protein Levels ◽  
Epithelial Membrane Protein 2 ◽  
Glycosylphosphatidyl Inositol ◽  
Reduced Surface

Caveolae are a subset of lipid rafts enriched in glycosphingolipids and cholesterol-rich domains, but selectively lacking glycosylphosphatidyl inositol-anchored proteins (GPI-APs). Caveolin proteins are the organizing component of caveolae, but the corresponding proteins for other classes of lipid rafts are poorly defined. Epithelial membrane protein-2 (EMP2), a member of the four-transmembrane superfamily, facilitates plasma membrane delivery of certain integrins. In this study, we found by laser confocal microscopy that EMP2 was associated with GPI-APs (detected by the GPI-AP binding bacterial toxin proaerolysin). Biochemical membrane fractionation and methyl-β-cyclodextrin treatment demonstrated that this association occurred within lipid rafts. EMP2 did not associate with caveolin-bearing membrane structures, and recombinant overexpression of EMP2 in NIH3T3 cells decreased caveolin-1 and caveolin-2 protein levels while increasing the surface expression of GPI-APs. Conversely, a ribozyme construct that specifically cleaves the EMP2 transcript reduced surface GPI-APs and increased caveolin protein expression. These findings suggest that EMP2 facilitates the formation and surface trafficking of lipid rafts bearing GPI-APs, and reduces caveolin expression, resulting in impaired formation of caveolae.

scholarly journals Feedback-Driven Mechanisms Between Phosphorylated Caveolin-1 and Contractile Actin Assemblies Instruct Persistent Cell Migration

2021 ◽  
Vol 9 ◽  
Author(s):  
Xuemeng Shi ◽  
Zeyu Wen ◽  
Yajun Wang ◽  
Yan-Jun Liu ◽  
Kun Shi ◽  
...  
Keyword(s):  
Cell Migration ◽  
Actin Filaments ◽  
Stress Fibers ◽  
Caveolin 1 ◽  
Contractile Stress ◽  
Active Processes ◽  
Outstanding Question ◽  
Directional Cell Migration ◽  
And Control ◽  
Persistent Cell

The actin cytoskeleton and membrane-associated caveolae contribute to active processes, such as cell morphogenesis and motility. How these two systems interact and control directional cell migration is an outstanding question but remains understudied. Here we identified a negative feedback between contractile actin assemblies and phosphorylated caveolin-1 (CAV-1) in migrating cells. Cytoplasmic CAV-1 vesicles display actin-associated motilities by sliding along actin filaments or/and coupling to do retrograde flow with actomyosin bundles. Inhibition of contractile stress fibers, but not Arp2/3-dependent branched actin filaments, diminished the phosphorylation of CAV-1 on site Tyr14, and resulted in substantially increased size and decreased motility of cytoplasmic CAV-1 vesicles. Reciprocally, both the CAV-1 phospho-deficient mutation on site Tyr14 and CAV-1 knockout resulted in dramatic AMPK phosphorylation, further causing reduced active level of RhoA-myosin II and increased active level of Rac1-PAK1-Cofilin, consequently led to disordered contractile stress fibers and prominent lamellipodia. As a result, cells displayed depolarized morphology and compromised directional migration. Collectively, we propose a model in which feedback-driven regulation between actin and CAV-1 instructs persistent cell migration.

scholarly journals Characterization of a novel virus infected watermelon in Indonesia based on viral particle using electron microscope

2017 ◽  
Author(s):  
Budi Setiadi Daryono ◽  
Fauziatul Fitriyah ◽  
alin liana ◽  
Utari Saraswati ◽  
Keiko T. Natsuaki
Keyword(s):  
Electron Microscope ◽  
Mosaic Virus ◽  
Viral Particle ◽  
Plant Viruses ◽  
Electron Microscope Investigation ◽  
Viral Particles ◽  
Cucumber Fruit ◽  
Novel Virus ◽  
New Form

Most of the cucurbits diseases in Indonesia are caused by plant viruses. Tobamovirus is one of the viral genera recently infected cucurbits. The members of cucurbits-infecting Tobamovirus are Cucumber green mottle mosaic virus (CGMMV), Kyuri green mottle mosaic virus (KGMMV), Cucumber fruit mottle mosaic virus (CFMMV), Zucchini green mottle mosaic virus (ZGMMV), and Cucumber mottle virus (CuMoV). A research on the occurrence of cucurbits-infecting Tobamovirus in Indonesia was carried out in 2011. Based on the result, a new characteristic of viral particle were found in watermelon. The viral particle was previously identified as CGMMV based on electron microscope investigation revealed presence of rod-shaped and not enveloped virions. The virions were straight about 300 nm long and 28–30 nm in diameter. However, in some electron micrograph shows rare characteristic of the viral particle. The viral particles could joint each other to bend a new form. Two viral particles were able to joint and formed an angle. Moreover, some of viral particles could joint and form a longer viral particle (800-1100 nm), two to four times longer than CGMMV particle, but the both diameter of the virus are similar. These characteristics indicate that the viral particle is a novel virus, different from CGMMV or other Tobamovirus members.

scholarly journals Adhesive Contact Between Cylindrical (Ebola) and Spherical (SARS-CoV-2) Viral Particles and a Cell Membrane

2020 ◽  
Author(s):  
Jiajun Wang ◽  
Nicole Lapinski ◽  
X. Frank Zhang ◽  
Anand Jagota
Keyword(s):  
Cell Membrane ◽  
External Force ◽  
Viral Particle ◽  
Bending Stiffness ◽  
Applied Force ◽  
Viral Particles ◽  
Contact Width ◽  
Initial Adhesion ◽  
A Cell ◽  
Force Contact

AbstractA critical event during the process of cell infection by a viral particle is attachment, which is driven by adhesive interactions and resisted by bending and tension. The biophysics of this process has been studied extensively but the additional role of externally applied force or displacement has generally been neglected. In this work we study the adhesive force-displacement response of viral particles against a cell membrane. We have built two models: one in which the viral particle is cylindrical (say, representative of filamentous virus such as Ebola) and another in which it is spherical (such as SARS-CoV-2 and Zika). Our interest is in initial adhesion, in which case deformations are small and the mathematical model for the system can be simplified considerably. The parameters that characterize the process combine into two dimensionless groups that represent normalized membrane bending stiffness and tension. In the limit where bending dominates, for sufficiently large values of normalized bending stiffness, there is no adhesion between viral particles and the cell membrane without applied force. (The zero-external-force contact width and pull-off force are both zero.) For large values of normalized membrane tension, the adhesion between virus and cell membrane is weak but stable. (The contact width at zero external force has a small value.) Our results for pull-off force and zero force contact width help to quantify conditions that could aid the development of therapies based on denying the virus entry into the cell by blocking its initial adhesion.

scholarly journals PostexitSurface Engineering of Retroviral/Lentiviral Vectors

2013 ◽  
Vol 2013 ◽  
pp. 1-8 ◽  
Author(s):  
Christoph Metzner ◽  
Feliks Kochan ◽  
John A. Dangerfield
Keyword(s):  
Chemical Modification ◽  
Genetic Information ◽  
Viral Particle ◽  
Genetic Manipulation ◽  
Regulatory Elements ◽  
Safe Delivery ◽  
Advantages And Disadvantages ◽  
Genomic Level ◽  
Gene Delivery Vectors ◽  
Viral Particles

Gene delivery vectors based on retroviral or lentiviral particles are considered powerful tools for biomedicine and biotechnology applications. Such vectors require modification at the genomic level in the form of rearrangements to allow introduction of desired genes and regulatory elements (genotypic modification) as well as engineering of the physical virus particle (phenotypic modification) in order to mediate efficient and safe delivery of the genetic information to the target cell nucleus. Phenotypic modifications are typically introduced at the genomic level through genetic manipulation of the virus producing cells. However, this paper focuses on methods which allow modification of viral particle surfaces after they have exited the cell, that is, directly on the viral particles in suspension. These methods fall into three categories: (i) direct covalent chemical modification, (ii) membrane-topic reagents, and (iii) adaptor systems. Current applications of such techniques will be introduced and their advantages and disadvantages will be discussed.

scholarly journals Nipah Virus-Like Particle Egress Is Modulated by Cytoskeletal and Vesicular Trafficking Pathways: a Validated Particle Proteomics Analysis

mSystems ◽  
2019 ◽  
Vol 4 (5) ◽  
Author(s):  
Gunner P. Johnston ◽  
Birgit Bradel-Tretheway ◽  
Paul D. Piehowski ◽  
Heather M. Brewer ◽  
Bom Nae Rin Lee ◽  
...  
Keyword(s):  
Viral Particle ◽  
Particle Production ◽  
Vaccine Development ◽  
Nipah Virus ◽  
Particle Formation ◽  
Vesicular Trafficking ◽  
Cellular Processes ◽  
Viral Particle Production ◽  
Viral Particles ◽  
Cellular Machinery

ABSTRACT Classified as a biosafety level 4 (BSL4) select agent, Nipah virus (NiV) is a deadly henipavirus in the Paramyxoviridae family, with a nearly 75% mortality rate in humans, underscoring its global and animal health importance. Elucidating the process of viral particle production in host cells is imperative both for targeted drug design and viral particle-based vaccine development. However, little is understood concerning the functions of cellular machinery in paramyxoviral and henipaviral assembly and budding. Recent studies showed evidence for the involvement of multiple NiV proteins in viral particle formation, in contrast to the mechanisms understood for several paramyxoviruses as being reliant on the matrix (M) protein alone. Further, the levels and purposes of cellular factor incorporation into viral particles are largely unexplored for the paramyxoviruses. To better understand the involvement of cellular machinery and the major structural viral fusion (F), attachment (G), and matrix (M) proteins, we performed proteomics analyses on virus-like particles (VLPs) produced from several combinations of these NiV proteins. Our findings indicate that NiV VLPs incorporate vesicular trafficking and actin cytoskeletal factors. The involvement of these biological processes was validated by experiments indicating that the perturbation of key factors in these cellular processes substantially modulated viral particle formation. These effects were most impacted for NiV-F-modulated viral particle formation either autonomously or in combination with other NiV proteins, indicating that NiV-F budding relies heavily on these cellular processes. These findings indicate a significant involvement of the NiV fusion protein, vesicular trafficking, and actin cytoskeletal processes in efficient viral particle formation. IMPORTANCE Nipah virus is a zoonotic biosafety level 4 agent with high mortality rates in humans. The genus to which Nipah virus belongs, Henipavirus, includes five officially recognized pathogens; however, over 20 species have been identified in multiple continents within the last several years. As there are still no vaccines or treatments for NiV infection, elucidating its process of viral particle production is imperative both for targeted drug design as well as for particle-based vaccine development. Developments in high-throughput technologies make proteomic analysis of isolated viral particles a highly insightful approach to understanding the life cycle of pathogens such as Nipah virus.

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