scholarly journals Polymorphism and Structural Maturation of Bunyamwera Virus in Golgi and Post-Golgi Compartments

2003 ◽  
Vol 77 (2) ◽  
pp. 1368-1381 ◽  
Author(s):  
Iñigo J. Salanueva ◽  
Reyes R. Novoa ◽  
Pilar Cabezas ◽  
Carmen López-Iglesias ◽  
José L. Carrascosa ◽  
...  
Keyword(s):  
Golgi Complex ◽  
Vero Cells ◽  
Productive Infection ◽  
Tubular Structures ◽  
Enveloped Viruses ◽  
Golgi Stack ◽  
Golgi Region ◽  
Viral Particles ◽  
Infected Cells ◽  
Bunyamwera Virus

ABSTRACT The Golgi apparatus is the assembly site for a number of complex enveloped viruses. Using high-preservation methods for electron microscopy, we have detected two previously unknown maturation steps in the morphogenesis of Bunyamwera virus in BHK-21 cells. The first maturation takes place inside the Golgi stack, where annular immature particles transform into dense, compact structures. Megalomicin, a drug that disrupts the trans side of the Golgi complex, reversibly blocks transformation, showing that a functional trans-Golgi is needed for maturation. The second structural change seems to take place during the egress of viral particles from cells, when a coat of round-shaped spikes becomes evident. A fourth viral assembly was detected in infected cells: rigid tubular structures assemble in the Golgi region early in infection and frequently connect with mitochondria. In Vero cells, the virus induces an early and spectacular fragmentation of intracellular membranes while productive infection progresses. Assembly occurs in fragmented Golgi stacks and generates tubular structures, as well as the three spherical viral forms. These results, together with our previous studies with nonrelated viruses, show that the Golgi complex contains key factors for the structural transformation of a number of enveloped viruses that assemble intracellularly.

scholarly journals The rubella virus E2 and E1 spike glycoproteins are targeted to the Golgi complex.

1993 ◽  
Vol 121 (2) ◽  
pp. 269-281 ◽  
Author(s):  
T C Hobman ◽  
L Woodward ◽  
M G Farquhar
Keyword(s):  
Membrane Proteins ◽  
Golgi Complex ◽  
Rubella Virus ◽  
Cell Types ◽  
Vero Cells ◽  
Type I ◽  
Surface Transport ◽  
Golgi Stack ◽  
Golgi Region ◽  
Different Cell Types

Rubella virus (RV) has been reported to bud from intracellular membranes in certain cell types. In this study the intracellular site of targeting of RV envelope E2 and E1 glycoproteins has been investigated in three different cell types (CHO, BHK-21 and Vero cells) transfected with a cDNA encoding the two glycoproteins. By indirect immunofluorescence, E2 and E1 were localized to the Golgi region of all three cell types, and their distribution was disrupted by treatment with BFA or nocodazole. Immunogold labeling demonstrated that E2 and E1 were localized to Golgi cisternae and indicated that the glycoproteins were distributed across the Golgi stack. Analysis of immunoprecipitates obtained from stably transfected CHO cells revealed that E2 and E1 become endo H resistant and undergo sialylation without being transported to the cell surface. Transport of RV glycoproteins to the Golgi complex was relatively slow (t1/2 = 60-90 min). Coprecipitation experiments indicated that E2 and E1 form a heterodimer in the RER. E1 was found to fold much more slowly than E2, suggesting that the delay in transport of the heterodimer to the Golgi may be due to the slow maturation of E1 in the ER. These results indicate that RV glycoproteins behave as integral membrane proteins of the Golgi complex and thus provide a useful model to study targeting and turnover of type I membrane proteins in this organelle.

Structural Maturation of the Transmissible Gastroenteritis Coronavirus

1999 ◽  
Vol 73 (10) ◽  
pp. 7952-7964 ◽  
Author(s):  
Iñigo J. Salanueva ◽  
José L. Carrascosa ◽  
Cristina Risco
Keyword(s):  
Golgi Complex ◽  
Partial Recovery ◽  
Extracellular Medium ◽  
Golgi Stack ◽  
Viral Particles ◽  
Infected Cells ◽  
Monensin A ◽  
Transmissible Gastroenteritis ◽  
Morphological Maturation ◽  
Transmissible Gastroenteritis Coronavirus

ABSTRACT During the life cycle of the transmissible gastroenteritis coronavirus (TGEV), two types of virus-related particles are detected in infected swine testis cells: large annular viruses and small dense viruses. We have studied the relationships between these two types of particles. Immunoelectron microscopy showed that they are closely related, since both large and small particles reacted equally with polyclonal and monoclonal antibodies specific for TGEV proteins. Monensin, a drug that selectively affects the Golgi complex, caused an accumulation of large annular viral particles in perinuclear elements of the endoplasmic reticulum-Golgi intermediate compartment. A partial reversion of the monensin blockade was obtained in both the absence and presence of cycloheximide, a drug that prevented the formation of new viral particles. After removal of monensin, the Golgi complex recovered its perinuclear location, and a decrease in the number of perinuclear large viral particles was observed. The release of small dense viral particles into secretory vesicles and the extracellular medium was also observed, as was a partial recovery of infectivity in culture supernatants. Small viral particles started to be seen between the third and the fourth Golgi cisternae of normally infected cells. All of these data strongly indicate that the large annular particles are the immature precursors of the small dense viruses, which are the infectious TGEV virions. The immature viral particles need to reach a particular location at the trans side of the Golgi stack to complete their morphological maturation.

scholarly journals Ultrastructural analysis of SARS-CoV-2 interactions with the host cell via high resolution scanning electron microscopy

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Lucio Ayres Caldas ◽  
Fabiana Avila Carneiro ◽  
Luiza Mendonça Higa ◽  
Fábio Luiz Monteiro ◽  
Gustavo Peixoto da Silva ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
High Resolution ◽  
Vero Cells ◽  
Viral Interactions ◽  
Viral Particles ◽  
Direct Cell ◽  
Infected Cells ◽  
Scanning Electron ◽  
Intercellular Connections

Abstract SARS-CoV-2 is the cause of the ongoing COVID-19 pandemic. Here, we investigated the interaction of this new coronavirus with Vero cells using high resolution scanning electron microscopy. Surface morphology, the interior of infected cells and the distribution of viral particles in both environments were observed 2 and 48 h after infection. We showed areas of viral processing, details of vacuole contents, and viral interactions with the cell surface. Intercellular connections were also approached, and viral particles were adhered to these extensions suggesting direct cell-to-cell transmission of SARS-CoV-2.

Electron Microscopic study of human herpesvirus-6 (HHV-6)

1990 ◽  
Vol 48 (3) ◽  
pp. 384-385
Author(s):  
Mariko Yoshida ◽  
Fumio Uno ◽  
Koichi Yamanishi ◽  
Kazuyoshi Ikuta ◽  
Takeshi Kurata ◽  
...  
Keyword(s):  
Human Lymphocytes ◽  
Surface Membrane ◽  
Human Herpesvirus ◽  
Viral Assembly ◽  
Tubular Structures ◽  
Electron Microscopic ◽  
Cytoplasmic Vacuoles ◽  
Exanthem Subitum ◽  
Viral Particles ◽  
Infected Cells

Human lymphocytes from cord blood and MT-4 cells were infected with the Hashimoto strain of HHV-6 isolated from a patient with exanthem subitum or with the Z29 strain of this virus isolated from a patient with AIDS, and the infected cells were examined by transmission and scanning electron microscopy. The following observations, characteristic for infection with HHV-6, were obtained. In the nucleus, most of the capsids with a diameter of about 100nm had cores of low density, which seemed to consist of punctate and/or filamentous structures. Tubular structures formed due to some mistakes in the viral assembly were also detected. The most striking characteristic of the ultra structure of this virus was the distinct coating of intracytop1asmic and extra cellular nucleocapsids with a tegument of moderate electron density. These particles coated with a tegument acquired their envelopes by budding at the vacuolar membrane in the cytoplasm. Enveloped viral particles with a diameter of about 170-200 nm were observed in cytoplasmic vacuoles and on the cell surface membrane (Fig. 1).

scholarly journals Effect of caffeine and reduced temperature (20 degrees C) on the organization of the pre-Golgi and the Golgi stack membranes.

1993 ◽  
Vol 120 (6) ◽  
pp. 1321-1335 ◽  
Author(s):  
J Jäntti ◽  
E Kuismanen
Keyword(s):  
Golgi Complex ◽  
Brefeldin A ◽  
Microscopic Analysis ◽  
Electron Microscopic ◽  
Transport Pathways ◽  
Golgi Stack ◽  
Golgi Membranes ◽  
Golgi Region ◽  
Functional Interface ◽  
Reduced Temperature

In the present study we have dissected the transport pathways between the ER and the Golgi complex using a recently introduced (Kuismanen, E., J. Jäntti, V. Mäkiranta, and M. Sariola. 1992. J. Cell Sci. 102:505-513) inhibition of transport by caffeine at 20 degrees C. Recovery of the Golgi complex from brefeldin A (BFA) treatment was inhibited by caffeine at reduced temperature (20 degrees C) suggesting that caffeine inhibits the membrane traffic between the ER and the Golgi complex. Caffeine at 20 degrees C did not inhibit the BFA-induced retrograde movement of the Golgi membranes. Further, incubation of the cells in 10 mM caffeine at 20 degrees C had profound effects on the distribution and the organization of the pre-Golgi and the Golgi stack membranes. Caffeine treatment at 20 degrees C resulted in a selective and reversible translocation of the pre- and cis-Golgi marker protein (p58) to the periphery of the cell. This caffeine-induced effect on the Golgi complex was different from that induced by BFA, since mannosidase II, a Golgi stack marker, remained perinuclearly located and the Golgi stack coat protein, beta-COP, was not detached from Golgi membranes in the presence of 10 mM caffeine at 20 degrees C. Electron microscopic analysis showed that, in the presence of caffeine at 20 degrees C, the morphology of the Golgi stack was altered and accumulation of numerous small vesicles in the Golgi region was observed. The results in the present study suggest that caffeine at reduced temperature (20 degrees C) reveals a functional interface between the pre-Golgi and the Golgi stack.

scholarly journals Beta-COP localizes mainly to the cis-Golgi side in exocrine pancreas.

1993 ◽  
Vol 121 (1) ◽  
pp. 49-59 ◽  
Author(s):  
A Oprins ◽  
R Duden ◽  
T E Kreis ◽  
H J Geuze ◽  
J W Slot
Keyword(s):  
Golgi Complex ◽  
Brefeldin A ◽  
Coated Vesicle ◽  
Aluminum Fluoride ◽  
Golgi Stack ◽  
Pancreatic Cells ◽  
Golgi Region ◽  
A Minor ◽  
Large Clusters ◽  
Small Clusters

We examined the distribution of the non-clathrin-coated vesicle-associated coat protein beta-COP in rat exocrine pancreatic cells by immunogold cytochemistry. Labeling for beta-COP was found in the Golgi region (48%) where it was associated with vesicles and buds of approximately 50 nm, showing a characteristic approximately 10-nm-thick coat. The other half of the label was present in the cytoplasm, not associated with visible coats or membranes, with a minor fraction present on small clusters of tubules and vesicles. Clathrin-coated vesicles were typically located at the trans-side of the Golgi complex, and showed a thicker coat of approximately 18 nm. Of the total beta-COP labeling over the Golgi region, 68% occurred on the cis-side, 6% on the cisternae, 17% on the rims of the cisternae, and only 9% on the trans-side. For clathrin these figures were 16, 2, 4, and 78%, respectively. At the cis-Golgi side beta-COP was present in transitional areas (TA), on so-called peripheral elements (PE), consisting of tubules and vesicles located between the cup-shaped transitional elements (TE) of the RER and the cis-most Golgi cisternae. Label for Sec23p was also present in TA but was located closer to the TE, while beta-COP labeled PE were located near the cis-Golgi cisternae. Upon energy depletion, Golgi associated beta-COP was almost exclusively (86%) in spherical aggregates of 200-500 nm in diameter, whereas the cis-side (6%), the cisternae (1%), the rims (4%) and trans-side (3%) of the Golgi complex, were barely labeled; 50% of the total label remained in the cytoplasm. The aggregates were predominantly located at the cis-side of the Golgi stack, next to, but distinct from the Sec23p positive TA, that were devoid of beta-COP and had only a few recognizable vesicles left. Incubation with aluminum fluoride resulted in fragmentation of the Golgi complex into large clusters of beta-COP positive vesicles, while 50% of the label remained in the cytoplasm, as in control cells. After 10 min of Brefeldin A treatment 91% of beta-COP was cytoplasmic and only 7% associated with membranes of the Golgi complex. The total label for beta-COP over exocrine cells remained unchanged during the incubation with either of the drugs, indicating that the drugs induce reallocation of beta-COP. Our data suggest that beta-COP plays a role in membrane transport at the cis-side of the Golgi complex.

scholarly journals Investigation of Extracellular Vesicles From SARS-CoV-2 Infected Specimens: A Safety Perspective

2021 ◽  
Vol 12 ◽  
Author(s):  
Yury O. Nunez Lopez ◽  
Anna Casu ◽  
Richard E. Pratley
Keyword(s):  
Disease Progression ◽  
Extracellular Vesicles ◽  
Research Needs ◽  
Scientific Communities ◽  
Safety Measures ◽  
Enveloped Viruses ◽  
Viral Particles ◽  
Viremic Phase ◽  
Infected Cells

The coronavirus disease 2019 (COVID-19) pandemic, caused by the SARS-CoV-2 virus, is wreaking havoc around the world. Considering that extracellular vesicles (EVs) released from SARS-CoV-2 infected cells might play a role in a viremic phase contributing to disease progression and that standard methods for EV isolation have been reported to co-isolate viral particles, we would like to recommend the use of heightened laboratory safety measures during the isolation of EVs derived from SARS-CoV-2 infected tissue and blood from COVID-19 patients. Research needs to be conducted to better understand the role of EVs in SARS-CoV-2 infectivity, disease progression, and transmission. EV isolation procedures should include approaches for protection from SARS-CoV-2 contamination. We recommend the EV and virology scientific communities develop collaborative projects where relationships between endogenous EVs and potentially lethal enveloped viruses are addressed to better understand the risks and pathobiology involved.

scholarly journals Ultrastructural analysis of SARS-CoV-2 interactions with the host cell via high resolution scanning electron microscopy

2020 ◽  
Author(s):  
Lucio Ayres Caldas ◽  
Fabiana Avila Carneiro ◽  
Luiza Mendonca Higa ◽  
Fabio Luiz Monteiro ◽  
Gustavo Peixoto da Silva ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
High Resolution ◽  
Vero Cells ◽  
Viral Interactions ◽  
Viral Particles ◽  
Direct Cell ◽  
Infected Cells ◽  
Scanning Electron ◽  
Intercellular Connections

SARS-CoV-2 is the cause of the ongoing COVID-19 pandemic. Here, we investigated the interaction of this new coronavirus with Vero cells using high resolution scanning electron microscopy. Surface morphology, the interior of infected cells and the distribution of viral particles in both environments were observed 2 and 48 hours after infection. We showed areas of viral processing, details of vacuole contents, and viral interactions with the cell surface. Intercellular connections were also approached, and viral particles were adhered to these extensions suggesting direct cell-to-cell transmission of SARS-CoV-2.

scholarly journals Key Golgi Factors for Structural and Functional Maturation of Bunyamwera Virus

2005 ◽  
Vol 79 (17) ◽  
pp. 10852-10863 ◽  
Author(s):  
Reyes R. Novoa ◽  
Gloria Calderita ◽  
Pilar Cabezas ◽  
Richard M. Elliott ◽  
Cristina Risco
Keyword(s):  
Conformational Changes ◽  
Secretory Pathway ◽  
Dependent Manner ◽  
Sugar Composition ◽  
Enveloped Viruses ◽  
Functional Maturation ◽  
Viral Particles ◽  
Viral Glycoproteins ◽  
Infected Cells ◽  
Viral Form

ABSTRACT Several complex enveloped viruses assemble in the membranes of the secretory pathway, such as the Golgi apparatus. Among them, bunyaviruses form immature viral particles that change their structure in a trans-Golgi-dependent manner. To identify key Golgi factors for viral structural maturation, we have purified and characterized the three viral forms assembled in infected cells, two intracellular intermediates and the extracellular mature virion. The first viral form is a pleomorphic structure with fully endo-β-N-acetylglucosaminidase H (Endo-H)-sensitive, nonsialylated glycoproteins. The second viral intermediate is a structure with hexagonal and pentagonal contours and partially Endo-H-resistant glycoproteins. Sialic acid is incorporated into the small glycoprotein of this second viral form. Growing the virus in glycosylation-deficient cells confirmed that acquisition of Endo-H resistance but not sialylation is critical for the trans-Golgi-dependent structural maturation and release of mature viruses. Conformational changes in viral glycoproteins triggered by changes in sugar composition would then induce the assembly of a compact viral particle of angular contours. These structures would be competent for the second maturation step, taking place during exit from cells, that originates fully infectious virions.

Epizootic hemorrhagic disease virus of deer: an electron microscopic study

1970 ◽  
Vol 16 (6) ◽  
pp. 427-432 ◽  
Author(s):  
Kin-Son Tsai ◽  
Lars Karstad
Keyword(s):  
Negative Contrast ◽  
Hemorrhagic Disease ◽  
Newborn Mouse ◽  
Tubular Structures ◽  
Electron Microscopic ◽  
Virus Particles ◽  
Dense Granules ◽  
Epizootic Hemorrhagic Disease ◽  
Viral Particles ◽  
Infected Cells

A mouse brain adapted strain of epizootic hemorrhagic disease (EHD) virus of deer was studied by ultrathin section and negative contrast electron microscopy. In sections of infected BHK-21 cells and neurons of newborn mouse brains, aggregates of virus particles, some appearing to be lacking their outer coats, were observed within membrane-bound vesicles. The development of virus particles was associated with intracytoplasmic viral matrices, consisting of moderately electron-dense granules. The viral particles measured about 58.8 nm in diameter in ultrathin sections and 62.3 nm in negative contrast preparations. Tubular structures, closely associated with viral matrices, were frequently found in infected cells. In one deer intracerebrally inoculated with virulent EHD virus, viral matrices, virus particles, and tubular structures, identical with those in infected BHK-21 cells and mouse neurons, were found in the cytoplasm of endothelial cells of blood vessels. The morphological details of EHD virus are compared with those of other viruses, and the differences in structure are discussed.

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