Morphological and physical properties of a multiploid-forming mutant of Western equine encephalitis virus (original) (raw)
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Specific membranous structures associated with the replication of group A arboviruses
Journal of virology, 1972
INTRACYTOPLASMIC MEMBRANOUS STRUCTURES OF A UNIQUE TYPE WERE ASSOCIATED WITH THE REPLICATION OF THREE GROUP A ARBOVIRUSES: Semliki Forest virus (SFV), Sindbis virus, or Western equine encephalomyelitis virus. The structures, referred to as type 1 cytopathic vacuoles (CPV-1), were membrane-limited and characteristically lined by regular membranous spherules measuring 50 nm in diameter. The membranous spherules typically contained a fine central density, but were neither virus cores nor virions. Detection of CPV-1 by electron microscopy at 3 to 6 hr postinfection coincided with the time of rapid virus growth and preceded the accumulation of virus nucleocapsids. A range of 20 to 100 CPV-1 profiles were counted per 100 ultrathin cell sections at 6 to 9 hr postinfection when viruses were grown in chick embryo, baby hamster kidney, or mouse L cells. Maximum counts remained in the same range even when the multiplicity of infection was varied over 100-fold. Inhibition of cellular ribonuclei...
1995
The formation of Hantaan (HTN) virus nucleocapsid-like structures (NLS) or virus-like particles (VLP) from expressed gene products was investigated in two eukaryotic systems. Baculovirus expression of the HTN virus small segment (S), which encodes the viral nucleocapsid protein, resulted in assembly of NLS inside infected insect cells. The NLS and authentic ribonucleocapsids, prepared by detergent disruption of HTN virions, had similar sedimentation characteristics and morphologies, and were recognized by HTN virus Nspecific antibodies. Co-expression of S and the medium segment (M), which encodes the two viral envelope glycoproteins (G1 and G2), did not efficiently generate VLP in the baculovirus-insect cell system, but VLP were observed in lysates and supernatants of cells infected with a recombinant vaccinia virus co-expressing HTN virus M and S. The VLP sedimented in sucrose to densities consistent with HTN virions, and some of them bore a striking resemblance to Hantaan virions when examined by immunoelectron microscopy.
Journal of Virology, 1977
We report here an in vitro system designed to study the interactions of vesicular stomatitis virus (VSV) proteins with cellular membranes. We have synthesized the VSV nucleocapsid (N) protein, nonstructural (NS) protein, glycoprotein (G protein), and membrane (M) protein in a wheat germ, cell-free, protein-synthesizing system directed by VSV 12 to 18S RNA. When incubated at low salt concentrations with purified cytoplasmic membranes derived from Chinese hamster ovary cells, the VSV M and G proteins bind to membranes, whereas the VSV N and NS proteins do not. The VSV M protein binds to membranes in low or high divalent cation concentrations, whereas binding of significant amounts of G protein requires at least 5 mM magnesium acetate concentrations. Vesicular stomatitis virus (VSV) is a simple, lar membranes, whereas the VSV N and NS enveloped virus that contains two membrane proteins do not. proteins: the glycoprotein (G protein), which forms the spikes of the virion (4, 23), and the MATERIALS AND METHODS membrane (M) protein, which lines the inner Cells and viruses. Membranes were prepared surface of the viral membrane (3). There are from CHO cells. Stocks of VSV (pure B particles of three other known viral proteins, the VSV nuthe Indiana serotype) were grown in CHO cells and cleocapsid (N) protein, the nonstructural (NS) purified as described previously (20). protein, and the viral transcriptase (L) protein. Preparation of VSV 12-186 polyribosomal RNA. These three proteins are associated with the The procedure described by Palmiter (15) was used core ofthe virus particle (16, 23, 24). Each of the with several modifications for the preparation of five viral proteins is synthesized from a mono-VSV 12-18S polyribosomal RNA. CHO cells growing cistronic mRNA (9, 13, 14). at 37°C were infected with VSV at a multiplicity of 3 risngtic m arNAl(9 13, 14). PFU/cell as described previously (20), except that 5 During the early stages in the maturation Of~.tg of actinomycin D per ml was added at the beginthis virus, host cell membranes are modified ning of infection. [3H]uridine (70 Ci/mmol, 25 ,uCi/ with the VSV G and M proteins. Cell fractionaml; New England Nuclear Corp.) was added 2 h tion studies of VSV-infected cells have shown postinfection. Infected cells were harvested at 4.5 h that the VSV M and G proteins rapidly become postinfection, suspended in sucrose-TKM buffer associated with the membrane fraction of the (0.05 M Tris [pH 7.5], 0.025 M KCl, 0.005 M magnecells after their synthesis (5, 11, 12, 24). Nucleo-sium acetate, 0.25 M sucrose), and disrupted with 10 capsid structures containing the genome RNA strokes of a tight-fitting Dounce homogenizer. Nuas well as the viral N, NS, and L proteins are clei were removed by centrifugation (1,000 x g for 2 asswemlld as the vira plaNsm. andsLptroteisures min). The resulting cytoplasmic extract was centriassembled in the cytoplasm. These structures fuged at 20,000 x g for 20 min, and the pellet (mem
Nucleocapsid and glycoprotein organization in an enveloped virus
Cell, 1995
Alphaviruses are a group of icosahedral, positive-strand RNA, enveloped viruses. The membrane bilayer, which surrounds the approximately 400 A diameter nucleocapsid, is penetrated by 80 spikes arranged in a T = 4 lattice. Each spike is a trimer of heterodimers consisting of glycoproteins E1 and E2. Cryoelectron microscopy and image reconstruction of Ross River virus showed that the T = 4 quaternary structure of the nucleocapsid consists of pentamer and hexamer clusters of the capsid protein, but not dimers, as have been observed in several crystallographic studies. The E1-E2 heterodimers form one-to-one associations with the nucleocapsid monomers across the lipid bilayer. Knowledge of the atomic structure of the capsid protein and our reconstruction allows us to identify capsid-protein residues that interact with the RNA, the glycoproteins, and adjacent capsid-proteins.
Structure and Composition of Viruses
Veterinary Virology, 1987
Structure and Composition of Viruses Morphology 4 Chemical Composition 9 Preservation of Viral Infectivity 18 Further Reading 19 The unicellular microorganisms can be arranged in order of decreas ing size and complexity: protozoa, fungi, bacteria, mycoplasmas, rickettsiae, and chlamydiae. These microorganisms, however small and simple, are cells. They always contain DNA as the repository of their genetic information, they contain RNA, and they have their own ma chinery for producing energy and macromolecules. Microorganisms grow by synthesizing their own macromolecular constituents (nucleic acid, protein, carbohydrate, and lipid), and they multiply by binary fission. Viruses, on the other hand, are smaller and simpler in construction than unicellular microorganisms, and they contain only one type of nucleic acid-either DNA or RNA, never both. Furthermore, since vi ruses have no ribosomes, mitochondria, or other organelles, they are completely dependent on their cellular hosts for energy production and protein synthesis. They replicate only within cells of the host that they infect. Indeed, unlike any microorganism, many viruses can, in suitable cells, reproduce themselves from their genome, a single nucleic acid molecule; i.e., their nucleic acid alone is infectious. Are viruses alive? The question is rhetorical. Outside a susceptible cell, the virus particle, 3 4 1. Structure and Composition of Viruses
Observations on the Structure of the Nucleocapsids of some Paramyxoviruses
Journal of General Virology, 1970
The intact particles and nucteocapsids of mumps, Sendai and measles viruses are of closely similar appearance, size and structure. The intact particles are about 15o nm. in diameter. The filamentous nucleocapsids have a modal length of about ~. I/Am., and are constructed of subunits arranged as a single start helix of pitch 5.0 nm. for Sendai virus, and about 6"o nm. for mumps and measles viruses. A (helical) projection of the structure of the Sendai nucleocapsid calculated from an electron micrograph showed that the structural subunits are hour-glass-shaped and are arranged in the helix with their long axes inclined at an angle of about 6o ° to the long axis of the particle. There are probably J ~ or I3 subunits in each turn of the basic helix. Optical diffraction patterns of electron micrographs of mumps and measles nucleocapsids show that they have closely similar structures.