H. Lodish - Molecular Cell Biology (5ed, Freeman, 2003) (796244), страница 57
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The host-cell range of some animal viruses is further restricted to a limited number of cell types because onlythese cells have appropriate surface receptors to which thevirions can attach.Viral Capsids Are Regular Arrays of Oneor a Few Types of ProteinThe nucleic acid of a virion is enclosed within a protein coat,or capsid, composed of multiple copies of one protein or afew different proteins, each of which is encoded by a singleviral gene.
Because of this structure, a virus is able to encodeall the information for making a relatively large capsid in asmall number of genes. This efficient use of genetic information is important, since only a limited amount of RNA orDNA, and therefore a limited number of genes, can fit intoa virion capsid. A capsid plus the enclosed nucleic acid iscalled a nucleocapsid.Nature has found two basic ways of arranging the multiple capsid protein subunits and the viral genome into a nucleocapsid. In some viruses, multiple copies of a single coatprotein form a helical structure that encloses and protects theviral RNA or DNA, which runs in a helical groove within theprotein tube. Viruses with such a helical nucleocapsid, suchas tobacco mosaic virus, have a rodlike shape.
The othermajor structural type is based on the icosahedron, a solid,approximately spherical object built of 20 identical faces,each of which is an equilateral triangle.The number and arrangement of coat proteins in icosahedral, or quasi-spherical, viruses differ somewhat depending on their size. In small viruses of this type, each of the 20triangular faces is constructed of three identical capsid protein subunits, making a total of 60 subunits per capsid.
Allthe protein subunits are in equivalent contact with one another (Figure 4-37a). In large quasi-spherical viruses, eachface of the icosahedron is composed of more than three subunits. As a result, the contacts between subunits not at thevertices are quasi-equivalent (Figure 4-37b). Models of several quasi-spherical viruses, based on cryoelectron microscopy, are shown in Figure 4-37. In the smaller viruses (e.g.,poliovirus), clefts that encircle each of the vertices of theicosahedral structure interact with receptors on the surfaceof host cells during infection. In the larger viruses (e.g., adenovirus), long fiberlike proteins extending from the nucleocapsid interact with cell-surface receptors on host cells.In many DNA bacteriophages, the viral DNA is locatedwithin an icosahedral “head” that is attached to a rodlike“tail.” During infection, viral proteins at the tip of the tailbind to host-cell receptors, and then the viral DNA passesdown the tail into the cytoplasm of the host cell.In some viruses, the symmetrically arranged nucleocapsid is covered by an external membrane, or envelope, which138CHAPTER 4 • Basic Molecular Genetic Mechanisms(a) Small icosahedral viruses(b) A large icosahedral virus2 153 45 1 243SV4010 nmPoliovirusCPMV▲ FIGURE 4-37 Structures of quasi-spherical (icosahedral)viruses.
The actual shape of the protein subunits in these virusesis not a flat triangle as illustrated in the schematic diagrams, butthe overall effect when the subunits are assembled is of a roughlyspherical structure with triangular faces. The three-dimensionalmodels are all shown at the same magnification. (a) In thesimplest and smallest quasi-spherical viruses, three identicalcapsid protein subunits form each triangular face (red) of theicosahedron (schematic).
The subunits meet in fivefold symmetryat each vertex. Models of three such viruses are shown: poliovirus,a human RNA virus; cowpea mosaic virus (CPMV), a plant RNAvirus; and simian virus 40 (SV40), a monkey DNA virus. (b) In somelarger viruses of this type, each triangular face is composed of sixsubunits. The subunits at the vertices maintain fivefold symmetry,but those making up the surfaces in between exhibit sixfoldsymmetry. A model of adenovirus, a human DNA virus, illustrateshow much larger it is than the viruses in part (a) and shows thefibers (green) that bind to receptors on host cells. [See P. L. StewartAdenovirus2136455 1 23et al., 1997, EMBO J. 16:1189. Models of CPMV, poliovirus, and SV40courtesy of T. S.
Baker; model of adenovirus courtesy of P. L. Stewart.]consists mainly of a phospholipid bilayer but also containsone or two types of virus-encoded glycoproteins (Figure4-38). The phospholipids in the viral envelope are similar tothose in the plasma membrane of an infected host cell. Theviral envelope is, in fact, derived by budding from that membrane, but contains mainly viral glycoproteins, as we discussshortly.Viruses Can Be Cloned and Countedin Plaque Assays▲ EXPERIMENTAL FIGURE 4-38 Viral protein spikesprotrude from the surface of an influenza virus virion.Influenza viruses are surrounded by an envelope consisting of aphospholipid bilayer and embedded viral proteins.
The large spikesseen in this electron micrograph of a negatively stained influenzavirion are composed of neuraminidase, a tetrameric protein, orhemagglutinin, a trimeric protein (see Figure 3-7). Inside is thehelical nucleocapsid. [Courtesy of A. Helenius and J. White.]The number of infectious viral particles in a sample can bequantified by a plaque assay.
This assay is performed by culturing a dilute sample of viral particles on a plate coveredwith host cells and then counting the number of local lesions, called plaques, that develop (Figure 4-39). A plaquedevelops on the plate wherever a single virion initially infects a single cell. The virus replicates in this initial host celland then lyses (ruptures) the cell, releasing many progenyvirions that infect the neighboring cells on the plate.
Aftera few such cycles of infection, enough cells are lysed to pro-4.7 • Viruses: Parasites of the Cellular Genetic System(a)Confluent layer of susceptible host cellsgrowing on surface of a plate139 EXPERIMENTAL FIGURE 4-39 Plaque assay determinesthe number of infectious particles in a viral suspension.(a) Each lesion, or plaque, which develops where a single virioninitially infected a single cell, constitutes a pure viral clone.(b) Plate illuminated from behind shows plaques formed bybacteriophage plated on E.
coli. (c) Plate showing plaquesproduced by poliovirus plated on HeLa cells. [Part (b) courtesy ofBarbara Morris; part (c) from S. E. Luria et al., 1978, General Virology,3d ed., Wiley, p. 26.]Add dilute suspension containing virus;after infection, cover layer of cellswith agar; incubate(b)Plaque(c)PlaquePlaqueEach plaque represents cell lysis initiated by one viralparticle (agar restricts movement so that virus caninfect only contiguous cells)duce a visible clear area, or plaque, in the layer of remaininguninfected cells.Since all the progeny virions in a plaque are derived froma single parental virus, they constitute a virus clone. Thistype of plaque assay is in standard use for bacterial and animal viruses.
Plant viruses can be assayed similarly by counting local lesions on plant leaves inoculated with viruses.Analysis of viral mutants, which are commonly isolated byplaque assays, has contributed extensively to current understanding of molecular cellular processes. The plaque assayalso is critical in isolating bacteriophage clones carryingsegments of cellular DNA, as discussed in Chapter 9.Lytic Viral Growth Cycles Lead to Deathof Host CellsAlthough details vary among different types of viruses, thosethat exhibit a lytic cycle of growth proceed through thefollowing general stages:1.
Adsorption—Virion interacts with a host cell by bindingof multiple copies of capsid protein to specific receptors onthe cell surface.2. Penetration—Viral genome crosses the plasma membrane.For animal and plant viruses, viral proteins also enter thehost cell.3. Replication—Viral mRNAs are produced with the aidof the host-cell transcription machinery (DNA viruses) orby viral enzymes (RNA viruses). For both types of viruses,viral mRNAs are translated by the host-cell translationmachinery. Production of multiple copies of the viralgenome is carried out either by viral proteins alone orwith the help of host-cell proteins.4. Assembly—Viral proteins and replicated genomesassociate to form progeny virions.5. Release—Infected cell either ruptures suddenly (lysis),releasing all the newly formed virions at once, or disintegrates gradually, with slow release of virions.Figure 4-40 illustrates the lytic cycle for T4 bacteriophage, a nonenveloped DNA virus that infects E.
coli. Viralcapsid proteins generally are made in large amounts becausemany copies of them are required for the assembly of eachprogeny virion. In each infected cell, about 100–200 T4progeny virions are produced and released by lysis.The lytic cycle is somewhat more complicated for DNAviruses that infect eukaryotic cells. In most such viruses, theDNA genome is transported (with some associated proteins)into the cell nucleus. Once inside the nucleus, the viral DNAis transcribed into RNA by the host’s transcription machinery. Processing of the viral RNA primary transcript by hostcell enzymes yields viral mRNA, which is transported to thecytoplasm and translated into viral proteins by host-cellribosomes, tRNA, and translation factors.
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