scholarly journals STRUCTURE OF TYPE 5 ADENOVIRUS

1963 ◽  
Vol 118 (2) ◽  
pp. 295-306 ◽  
Author(s):  
Wesley C. Wilcox ◽  
Harold S. Ginsberg
Keyword(s):  
Virus Particle ◽  
Density Gradient ◽  
Infectious Virus ◽  
Relative Proportion ◽  
Specific Antigen ◽  
Equilibrium Density ◽  
Virus Particles ◽  
Infected Cells ◽  
Specific Antigens ◽  
Virus Antigens

Type 5 adenovirus was purified by fluorocarbon (freon 113) treatment followed by banding in a CsCl equilibrium density gradient. This method permitted separation of virus from normal host cell materials and virus-specific soluble antigens. Virus banded in CsCl with a mean bouyant density of 1.3349 gm/cm3. The three virus-specific soluble antigens (group- and type-specific antigens and toxin) banded together with a mean bouyant density of 1.2832 gm/cm3. The group-specific antigen was the predominant antigen of the purified virus particle, whereas the group- and type-specific antigens were present in equal titers in the antigen band. Infectious virus particles were inactivated by prolonged dialysis at pH 10.5. Centrifugation of inactivated virus preparations in a CsCl equilibrium density gradient resulted in separation of virus DNA from specific antigen: the antigens banded with a mean bouyant density of 1.2832 gm/cm3 and the DNA sedimented to the bottom of the tube. The predominant antigen derived from purified virus particles was the group-specific antigen and it was in the same relative proportion to the type-specific antigen as measured in intact particles. The antigens derived from disrupted virus were immunologically identical with the soluble virus antigens present in infected cells.

scholarly journals Lösliche und virusgebundene Adenovirus Hämagglutinine

1964 ◽  
Vol 19 (7) ◽  
pp. 587-592 ◽  
Author(s):  
H. Bauer ◽  
R. Wigand ◽  
W. Adam
Keyword(s):  
Red Blood Cells ◽  
Density Gradient ◽  
Blood Cells ◽  
Infectious Virus ◽  
Cesium Chloride ◽  
Human Cells ◽  
Density Gradient Ultracentrifugation ◽  
Rat Blood ◽  
Virus Particles ◽  
Human Red Blood Cells

Prototype adenoviruses and their hemagglutinins of ROSEN'S 1 group II were studied by adsorption to rat and human red blood cells and by straight and cesium chloride density gradient ultracentrifugation. Density gradient experiments showed the presence of a hemagglutinin separable from the infectious virus particles which agglutinates both rat and human cells for virus types 9, 10, 13, 19, 26, and 27. In addition, types 10, 19, and 27 have a second hemagglutinin associated with the infective particles which agglutinates rat blood cells only. Type 9 (and possibly type 8) virus has a virusbound hemagglutinin which agglutinates both kinds of blood cells. The viruses types 15, 17, 22, and 24 with hemagglutinins for rat blood cells only have two hemagglutinins, one of them associated with the virus particles. The results with type 23 were variable. Straight ultracentrifugation experiments showed a surprisingly incomplete sedimentation of the infective particles for most of the virus types studied.Differences in the resistance of various hemagglutinins to trypsin and/or heating are demonstrated.

scholarly journals SYNTHESIS OF VIRUS-SPECIFIC POLYMERS IN ADENOVIRUS-INFECTED CELLS: EFFECT OF 5-FLUORODEOXYURIDINE

1962 ◽  
Vol 116 (2) ◽  
pp. 141-157 ◽  
Author(s):  
John F. Flanagan ◽  
Harold S. Ginsberg
Keyword(s):  
Hela Cells ◽  
Culture Medium ◽  
Deoxyribonucleic Acid ◽  
Infectious Virus ◽  
Potent Inhibitor ◽  
Virus Production ◽  
Inhibitory Effect ◽  
Virus Particles ◽  
Specific Proteins ◽  
Infected Cells

Biochemical synthesis in adenovirus-infected HeLa cells was studied utilizing 5-fluorodeoxyuridine (5-FUDR), a potent inhibitor of deoxyribonucleic acid production. Synthesis of saline-soluble DNA and infectious virus was completely suppressed by addition of the analogue to cells as late as 10 hours after infection. The inhibitory effect of this compound was totally reversed by addition of 10–6 M thymidine to the culture medium. Synthesis of DNA essential for virus production began 10 hours after infection and was completed by 16 hours after infection. These data support the hypothesis that the saline-soluble DNA is a precursor of infectious virus particles. Studies of antigen production indicated that formation of virus-specific proteins was directly dependent upon synthesis of DNA.

scholarly journals Coordination of Zika Virus Infection and Viroplasm Organization by Microtubules and Microtubule-Organizing Centers

Cells ◽  
2021 ◽  
Vol 10 (12) ◽  
pp. 3335
Author(s):  
Rebecca A. Buchwalter ◽  
Sarah C. Ogden ◽  
Sara B. York ◽  
Li Sun ◽  
Chunfeng Zheng ◽  
...  
Keyword(s):  
Zika Virus ◽  
Infectious Virus ◽  
Virus Production ◽  
Health Concern ◽  
Zikv Infection ◽  
Virus Particles ◽  
Microtubule Organizing Centers ◽  
Infected Cells ◽  
Congenital Microcephaly ◽  
A Site

Zika virus (ZIKV) became a global health concern in 2016 due to its links to congenital microcephaly and other birth defects. Flaviviruses, including ZIKV, reorganize the endoplasmic reticulum (ER) to form a viroplasm, a compartment where virus particles are assembled. Microtubules (MTs) and microtubule-organizing centers (MTOCs) coordinate structural and trafficking functions in the cell, and MTs also support replication of flaviviruses. Here we investigated the roles of MTs and the cell’s MTOCs on ZIKV viroplasm organization and virus production. We show that a toroidal-shaped viroplasm forms upon ZIKV infection, and MTs are organized at the viroplasm core and surrounding the viroplasm. We show that MTs are necessary for viroplasm organization and impact infectious virus production. In addition, the centrosome and the Golgi MTOC are closely associated with the viroplasm, and the centrosome coordinates the organization of the ZIKV viroplasm toroidal structure. Surprisingly, viroplasm formation and virus production are not significantly impaired when infected cells have no centrosomes and impaired Golgi MTOC, and we show that MTs are anchored to the viroplasm surface in these cells. We propose that the viroplasm is a site of MT organization, and the MTs organized at the viroplasm are sufficient for efficient virus production.

scholarly journals Mutations in the Influenza A Virus M1 Protein Enhance Virus Budding To Complement Lethal Mutations in the M2 Cytoplasmic Tail

2017 ◽  
Vol 92 (1) ◽  
Author(s):  
Hsuan Liu ◽  
Michael L. Grantham ◽  
Andrew Pekosz
Keyword(s):  
Virus Particle ◽  
Influenza A Virus ◽  
Influenza A ◽  
Infectious Virus ◽  
Virus Assembly ◽  
Virus Production ◽  
Cytoplasmic Tail ◽  
Filament Formation ◽  
Virus Budding ◽  
Virus Particles

ABSTRACTThe influenza A virus M1 and M2 proteins play important roles in virus assembly and in the morphology of virus particles. Mutations in the distal cytoplasmic tail region of M2, and in particular a tyrosine-to-alanine mutation at residue 76 (Y76A), were essential for infectious virus production and filament formation while having limited effects on total virus particle budding. Using a novel selection method, mutations at seven different M1 amino acids (residue 73, 94, 135, 136, or 138 or a double mutation, 93/244) that are not found in circulating influenza virus strains or have not been previously identified to play a role in influenza A virus assembly were found to complement the lethal M2Y76A mutation. These M1 suppressor mutations restored infectious virus production in the presence of M2Y76A and mediated increased budding and filament formation even in the absence of M2. However, the efficiency of infectious virus replication was still dependent on the presence of the distal region of the M2 cytoplasmic tail. The data suggest that influenza A virus budding and genome incorporation can occur independently and provide further support for complementary roles of the M1 and M2 proteins in virus assembly.IMPORTANCEInfluenza virus particle assembly involves the careful coordination of various viral and host factors to optimally produce infectious virus particles. We have previously identified a mutation at position 76 of the influenza A virus M2 protein that drastically reduces infectious virus production and filament formation with minimal effects on virus budding. In this work, we identified suppressor mutations in the M1 protein which complement this lethal M2 mutation by increasing the efficiency with which virus particles bud from infected cells and restoring filament formation at the infected-cell surface. M2 distal cytoplasmic domain sequences were still required for optimal infectivity. This indicates that M1 and M2 can functionally replace each other in some, but not all, aspects of virus particle assembly.

scholarly journals Mutant Human Cytomegalovirus Lacking the Immediate-Early TRS1 Coding Region Exhibits a Late Defect

2002 ◽  
Vol 76 (23) ◽  
pp. 12290-12299 ◽  
Author(s):  
Catherine A. Blankenship ◽  
Thomas Shenk
Keyword(s):  
Human Cytomegalovirus ◽  
Infectious Virus ◽  
Human Fibroblasts ◽  
Reporter Genes ◽  
Immediate Early ◽  
Viral Gene ◽  
Wild Type ◽  
Viral Mrnas ◽  
Virus Particles ◽  
Infected Cells

ABSTRACT The human cytomegalovirus IRS1 and TRS1 open reading frames encode immediate-early proteins with identical N-terminal domains and divergent C-terminal regions. Both proteins have been shown previously to activate reporter genes in transfection assays in cooperation with other viral gene products. We have constructed two viruses carrying substitution mutations within either the IRS1 or TRS1 open reading frame. ADsubIRS1 failed to produce the related IRS1 and IRS1263 proteins, but it replicated with normal kinetics to produce a wild-type yield in human fibroblasts. The addition in trans of the IRS1263 protein, which antagonizes the ability of IRS1 and TRS1 proteins to activate reporter genes, did not inhibit the growth of the mutant virus. ADsubTRS1 failed to produce the TRS1 protein, and it generated an ∼200-fold-reduced yield of infectious virus in comparison to its wild-type parent. Viral DNA accumulated normally, as did a set of viral mRNAs that were monitored in ADsubTRS1-infected cells. However, two tegument proteins were partially mislocalized and infectious virus particles did not accumulate to normal levels within ADsubTRS1-infected cells. Further, infectious ADsubTRS1 particles sedimented abnormally in a glycerol-tartrate gradient, indicating that the structure of the mutant particles is aberrant. Our analysis of the ADsubTRS1 phenotype indicates that the TRS1 protein is required, either directly or indirectly, for efficient assembly of virus particles.

scholarly journals Major Tegument Protein pp65 of Human Cytomegalovirus Is Required for the Incorporation of pUL69 and pUL97 into the Virus Particle and for Viral Growth in Macrophages

2008 ◽  
Vol 83 (6) ◽  
pp. 2480-2490 ◽  
Author(s):  
Meike Chevillotte ◽  
Sandra Landwehr ◽  
Leonhard Linta ◽  
Giada Frascaroli ◽  
Anke Lüske ◽  
...  
Keyword(s):  
Virus Particle ◽  
Human Cytomegalovirus ◽  
Cell Types ◽  
Viral Proteins ◽  
Tegument Protein ◽  
Viral Growth ◽  
Virus Particles ◽  
Infected Cells ◽  
Different Cell Types

ABSTRACT The tegument protein pp65 of human cytomegalovirus (HCMV) represents the major component of mature virus particles. Nevertheless, deletion of pp65 has been shown to have no effects on virus replication and morphogenesis in fibroblasts in vitro. We have studied the HCMV virion composition in the absence of pp65 and viral growth of a pp65 stop mutant in different cell types, including monocyte-derived macrophages. Two stop codons at amino acids 11 and 12 of pp65 were introduced by bacterial artificial chromosome mutagenesis into the endotheliotropic strain TB40/E. Clear changes of the tegument composition could be observed in purified mutant virus particles, where the amount of tegument protein pUL25 was drastically reduced. In addition, pUL69 and the virally encoded protein kinase UL97 were undetectable in the pp65 stop mutant. Expression of pUL69 in infected cells was unaltered while pUL25 accumulated in the absence of pp65, thus demonstrating that only incorporation into virus particles is dependent on pp65. Coimmunoprecipitation experiments using lysates of infected cells revealed an interaction between pUL69 and pp65. This interaction was verified in pull-down experiments using transfected cells, which showed that pp65 and pUL69 do not require the presence of other viral proteins for their interaction. We conclude that pp65 is required for the incorporation of other viral proteins into the virus particle and thus is involved in the protein-protein interaction network leading to normal tegument formation. When studying growth kinetics of the pp65 stop mutant in different cell types, we found a severe impairment of viral growth in monocyte-derived macrophages, showing for the first time a strong cell-specific role of pp65 in viral growth.

scholarly journals INCOMPLETE SIMIAN PAPOVAVIRUS SV40

1964 ◽  
Vol 119 (2) ◽  
pp. 313-326 ◽  
Author(s):  
Joseph L. Melnick ◽  
Sara E. Stinebaugh ◽  
Fred Rapp
Keyword(s):  
Viral Protein ◽  
Infectious Virus ◽  
Fluorescent Antibody ◽  
Virus Antigen ◽  
Fluorescent Antibody Technique ◽  
Antibody Technique ◽  
Antigenic Protein ◽  
Virus Particles ◽  
Large Numbers ◽  
Infected Cells

A study was made of the effects of 5-fluorouracil (FU) and 5-fluorodeoxyuridine (FUDR) on the replication of the simian papovavirus SV40 in cercopithecus monkey kidney cells and on the production of virus antigen by these cells. Both drugs markedly suppressed the production of new infectious virus by SV40-infected cells. Synthesis of viral protein was also markedly suppressed by FUDR, but not by FU. In the presence of FU, infected cells produced large amounts of viral protein which were detected by the fluorescent antibody technique. The antigen was not distributed in a particulate fashion as in untreated cells. Diffuse virus antigen was observed in the nuclei of FU-treated cells, resembling the distribution of antigen near the end of the eclipse period in untreated, infected cultures. This stage of antigen production presumably preceded viral assembly. Virus particles with or without cores were rarely seen with the electron microscope in infected FU-treated cells, although large numbers of SV40 particles were readily visualized in untreated, infected cells. It appears that at least one antigenic protein of this papovavirus is synthesized abundantly in FU-treated cells, but is not assembled into virus shells in the presence of the inhibitor.

scholarly journals STUDIES ON PERSISTENT INFECTIONS OF TISSUE CULTURES

1964 ◽  
Vol 119 (6) ◽  
pp. 895-922 ◽  
Author(s):  
José E. Rodriguez ◽  
Werner Henle
Keyword(s):  
Infectious Virus ◽  
Cellular Growth ◽  
Infectious Process ◽  
Virus Particles ◽  
Secondary Spread ◽  
Persistent Infections ◽  
Induce Resistance ◽  
Infected Cells ◽  
Vesicular Stomatitis ◽  
Infectious Cycle

The initial stages of infection of L(MCN) cell populations with standard Newcastle disease virus (NDVST) were analyzed in an effort to elucidate the steps leading to survival of the cultures and to the indefinite persistence of the infectious process at a low level. Cells were exposed in suspension to NDV at varying multiplicities and the monolayer cultures derived from such cells assayed at intervals for cellular growth rates, percentage of infected cells as determined by immunofluorescence, yields of viral progeny and of interferon, and, on occasion, resistance to superinfection with vesicular stomatitis virus. The percentage of cells calculated to be initially infected on the basis of adsorption data was found to match closely the percentage of immunofluorescent cells resulting from the first infectious cycle (up to 24 hours). Cells initially infected with NDVST produced a mixed progeny of infectious virus (from 15 to 40 pfu/cell) and about 10 times as many non-infectious particles in 24 hours [NDVL(MCN)], but little or no interferon. If all cells were infected the cultures ultimately died. At multiplicities of infection (m) of 2 or less the cultures survived with increasing ease as the percentage of infected cells was reduced. The number of pfu per infected cell was of the above order during the first 3 days; it declined thereafter. Limited secondary spread of the infection was noted by 48 hours and no further cycling was noted thereafter. As m decreased from 2.0 to 0.1 there was an increase in the yields of interferon and the time at which peak titers were reached. Addition of anti-NDV serum 2 hours after infection prevented measurable production of interferon. In contrast, following exposure of cells to NDVL(MCN) at multiplicities ranging from 20.0 to 0.2 (based on infectious virus) all cultures survived, no secondary spread was noted, the number of pfu per infected cells was reduced at the higher multiplicities, and the yields of interferon were similar and maximal by 24 hours and not affected by anti-NDV serum added after an adsorption period of 2 hours. It is concluded that the non-infectious virus particles in the progeny released from NDVST-infected cells induce resistance in remaining cells or, if adsorbed simultaneously with infectious virus, abort the intracellular infectious process. In both instances interferon is produced which may then render additional cells resistant. The non-infectious component is considered an incomplete or defective product of viral replication and not merely thermally inactivated virus. NDVST partially or completely inactivated at 37°C induced neither cellular resistance nor synthesis of interferon. The incomplete viral component behaved in all respects like ultraviolet-inactivated NDVST except that it was significantly more efficient in inducing interferon synthesis. On the basis of the presented data a scheme has been devised and discussed which appears to explain satisfactorily the events which take place on initial infection of L(MCN) cells with NDV and which lead to the persistence of the infectious process.

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