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Unformatted text preview: Structure and Classification of Mammalian Viruses Components of enveloped and naked viruses Nomenclature and classification of mammalian viruses How Viruses Infect Mammalian Cells and Replicate Themselves Overview of the steps in viral replication Influenza virus as an example of virus—host cell interactions Antiviral Compounds Pathology of Viral Disease Explaining symptoms Outcomes ofa viral infection Why Aren't We Doing Better? Viruses of Mammalian Cells i So, Naturalists observe, a flea has smaller fleas that on him prey,- and these have smaller still to bite ’em; and so proceed ad infinitam. —Ionathan Swift that all free-living cells have viruses capable of infecting them. The viruses of prokaryotic cells have been described in previous chap- ters. The viruses of eukaryotic microbes have been poorly characterized but clearly exist; plants and arthropods have a variety of viruses. in this Chapter, the focus will be on viruses of mammalian cells, the best characterized of all viruses of eukaryotic cells. This does not mean that viruses of plants, insects, and eukaryotic microbes are unimportant. In fact, the study of these viruses may well be a major area of research in the future. But, bowing to the reality of the huge literature on mammalian viruses, which may prove to be a valuable guide to future investigations of viruses of other eukaryotic cells, this chapter will confine itself to mammalian cell viruses, with an emphasis on viruses that attack human cells. In later chapters, examples of specific human viruses, such as hepatitis virus, polio virus, and human immunodeficiency virus (HIV), will be covered in more detail. This Chapter will serve as an introduction to the properties of such viruses. Viruses are totally dependent on free-living cells. In fact, it is likely Structure and Classification of Mammalian Viruses Components ofenveloped and naked viruses An introduction to viral structures was provided in Chapter 1 and will be reviewed only briefly here. The portion of a virus that is common to all viruses consists of a nucleic acid-protein core covered by a protein coat (capsid; Fig. 10.1). The capsid consists of many copies of one or a few polypeptides (capsomeres), which are tightly packed in an array that often has a crystalline appearance. The capsid gives the virus its shape, helps to organize the viral genome (which is packed inside it), and protects the viral genome from enzymes that might degrade it in the external environment. As will be seen shortly, the capsid also plays a role in the uncoating of the virus, the process by which the virus releases its genetic material into the I9?r I 98 Chapter I 0 Figure l0.l Components of naked and enveloped viruses. The nucleow protein core contams the viral genome, together with proteins that play a struc- tural role to organize the genome or per- form an enzymatic function during replication. The capsid consists of tightly packed proteins (capsomeres) that give the virus its shape and protect the nucleoprotein core. Enveloped viruses have a phospholipid bilayer membrane that contains envelope pro- teins, which are used for attachment to host cells. Underlying the envelope are matrix proteins that stabilize the mem- brane and attach it to the capsid. Capsid Nucleoprotein core Viral capsid Naked virus Phospholipid bilayer (derived from host cell) Capsid Nucleoprotein core Enveloped virus cell it is attacking. Later in the replication cycle, the capsid helps to orga- nize copies of the viral genome that have been synthesized inside the infected cell. Many viruses have an envelope, a protein-phospholipid layer that cov- ers the capsid. Viral envelopes contain proteins that project outward from the viral surface, envelope glycoproteins. Envelope glycoproteins mediate attachment of an enveloped virus to the host cell surface. They do this by binding very specifically to carbohydrates or proteins on the surfaces of the cells the virus infects. The host cell proteins bound by the viral surface pro- teins are sometimes called viral receptors. Matrix proteins, which interact with the membrane—embedded portion of the envelope protein and with capsid proteins, link the envelope to the capsid and help to stabilize the viral particle. Matrix proteins also play an important role in the uncoating of enveloped viruses and in the assembly of viral particles in late stages of the viral replication process. Viruses without an envelope are called unen- veloped or naked viruses. Because naked viruses have no envelopes, their capsid proteins are the ones that mediate attachment of the virus to the receptor molecules on the surface of the mammalian cell. As with enveloped viruses, the interaction between the capsid protein of the naked virus and the host cell viral receptor is very specific. Both naked and enveloped viruses have nucleoprotein cores enclosed in their capsids. The proteins of the nucleoprotein core can have a structural function, such as stabilization of the viral nucleic acid during initial and late stages of viral replication, or an enzymatic function. Depending on the way a virus replicates, it may have to provide some proteins that are not found in human cells. For example, the replication cycle of HIV involves a step in which a DNA copy is made of the RNA genome of the virus. Because mam- malian cells do not have enzymes that carry out such a reaction (reverse transcriptase), the virus must provide its own reverse transcriptase. Other examples of specialized replication enzymes provided by the virus will be given in a later section on strategies of viral replication. in contrast to free-living microbes, which all have double~stranded DNA genomes, viruses can have genomes composed of single-stranded RNA, double-stranded RNA, single—stranded DNA, or double-stranded DNA. A similar range of genome types is also seen in bacteria] viruses. Why viruses exhibit such a variety of genome types is not clear, but the fact that some of them have RNA genomes has a special significance for their evolution. RNA is less stable chemically than DNA. Moreover, enzymes that repro~ duce RNA tend to make more mistakes than those that reproduCe DNA. As a result, viruses with RNA genomes tend to mutate more rapidly than organisms with DNA genomes. Two viruses with RNA genomes, HIV and influenza virus, are among the most rapidly mutating organisms known. Not all RNA viruses mutate rapidly, however, so more than the RNA com- position of the viral genome is involved in viral mutation rates. Viral genomes range in size from about 4 kbp, the size of a small bac- terial plasmid, to about 200 kbp. The smallest bacterial genome is about 500 kbp in size. Thus, viruses range in genetic capacity from very simple, stripped-down genomes to complexities near that of a stripped-dOWn bac- terial genome. Viral genomes can be very small, because viruses have evolved to make use of many features of the cell they infect: the cell’s ability to synthesize nucleotides and amino acids, its ability to generate energy, and its ability to synthesize proteins and nucleic acids. Enveloped viruses even get their phosPholipid bilayer envelope from the nuclear membrane, the endoplasmic reticulum, or the cytoplasmic membrane of the infected cell. Thus, a virus needs only to supply genes encoding proteins of the enve- lope, capsid, and nucleoprotein core. The small genome is advantageous. A small genome can be replicated much more rapidly than a large one, allow- ing many copies of the viral genome to be made in a short period of time. In this way, viruses quickly outpace replication of the genome of the infected cell, competing successfully for components that should have been directed toward mammalian genome replication. Nomenclature and classification of mammalian viruses Virus classification is based on a number of characteristics (Table 10.1), including the presence or absence of an envelope, the shape of the virus, the characteristics of the virus’s capsomeres, the composition of its genome, and the characteristics of its replication strategy. In some cases, the host range of the virus and its mode of spread (by insect, for example) play a role in classification. Viruses are organized into families, genera, and species. The family name ends in “viridae” and is not italicized. The genus name, which is italicized, ends in “virus.” The species name, which is not italicized, also ends in “virus.” For example, the virus that causes herpes cold sores has the species name herpes simplex 1 virus and is a member of the genus Simplexviras and the family Herpesviridae. In this chapter, for Table In. I Characteristics used to classify mammalian viruses “— Presence or absence of envelope Shape of virus (e.g.. icosahedral.fi|amentous) Characteristics of viral genome RNA or DNA, single-stranded or double-stranded. linear or circular. segmented or nonsegmented Mode of spread (eg. insect vector) Viruses of Mammalian Cells I99 200 Chapter 10 simplicity, we will use primarily the family name and the species name. Examples of families of viruses covered in later chapters are provided in Figure 10.2. Evident relationships link virus families together taxonomically, as seen in Figure 10.2, but no single genetic yardstick, comparable to that for prokaryotic and eukaryofic ribosomal RNA sequences, exists for establishing relationships between mammalian viruses or between mammalian viruses and viruses of plants, arthropods, eukaryotic microbes, or prokaryotes. Figure [0.2 Examples of viral classification. All of the viruses included in this figure are described in later sections of the text. Note the use of features listed in Table 10.1 to organize the various families and species of viruses. For simplicity, only the family name and the species name are given in this figure. ssRNA genome ssRNA genome (+5trand) (—strand) Enveloped Nonsegmented genome. Dipioid genome. icosahedral capsid (DNA intermediate} l Nonsegmented genome Segmented genome Naked Enveioped EnveloPed. | coniczil capsid Picomaviridae Caliciviridae Fiaviviridoe Retroviri'doe Rhabdow'ridae Orthomyxaviridae Rhinovirus Norwalk virus ' (Chap. I6) (Chap. [7} Dengue virus HIV (Chap. IS) Rabies virus Influenza virus @ (Chap. 20) (Chap. l4) (Chaps. lo. l4. l6) Polio virus - (Chap. l5) .. HepatitisA virus (Chap. l7) ® dsRNA genome dsDNA genome Naked. icosahedral capsid. Icosahedral capsid segmented genome | Naked EnveloF-ed Reavilridae Popovoviridoe Hepodnaviridoe Herpesviridae nonvirus Papilioma virus Hepatitis B virus Herpes simplex viruses (Chap. l7) (Chap. 18) (Chap. l8) (Chap. :3) a a How Viruses Infect Mammalian Cells and Replicate Themselves Overview of the steps in viral replication The initial steps in the viral infection cycle are similar for all viruses. That is, the virus first binds to a Specific molecule on the surface of the cell it will infect (attachment), then releases its nucleoprotein core into the host cell cytoplasm, a process called uncoating. Some viruses follow attachment with a step that fuses their surfaces with the membrane of the target cell and allows release of the nucleoprotein core into the cell cytoplasm (Fig. 10.3). Others undergo a more complex internalization and uncoating process, which involves endocytosis of the virus, acidification of the endo» cytic vesicle interior, fusion of the viral surface to the vesicle membrane, and, finally, release of the nucleoprotein core into the host cell cytoplasm. After attachment and uncoating, copies of the viral genome and viral proteins are made. Viral genome replication can occur in the nucleus or in the cytOplasm, depending on the virus’s replication strategy. Replication strategies vary with the nature of the viral genome, as will be explained later in more detail. Viral proteins are synthesized in the cytOplasm. Finally, new viral genomes and viral proteins are assembled and exit the cell. The two main routes of exit are budding, a process by which the virus pushes through a membrane acquiring an envelope in the process, and lysis of the host cell cytoplasmic membrane, allowing viral exit without budding. Enveloped viruses bud through membranes of the infected Cell (nuclear membrane, endoplasmic reticulum membrane, or cytoplasmic membrane, depending on the virus). Simple as they seem at first glance, viruses exhibit an astonishing variety of strategies at each step of the repli- cation cycle. Influenza virus as an example of virus—host cell interactions To convey a sense of the complex interaction between a virus and a mam- malian cell, we will consider in detail a single type of virus, influenza virus. Influenza virus is not only a well-studied virus but it is also all too familiar to most people from personal experience with influenza, the disease caused by influenza virus. Thus, influenza virus provides a good illustration of virus—mammalian cell interaction. As we move through the stages of the influenza virus replication cycle, differences in the strategies used by other viruses will be noted. Another good reason to use influenza virus as an example is that, whereas influenza virus has some features in common with other viruses, it also has features that are unique. Its unique features under- score the fact that there is no such thing as a “typical” virus. Given their small sizes and relatively simple genomes, viruses display an impressive amount of variation in the ways they carry out different steps in their repli- cation cycle, Attachment and uncoating. Viruses seem so simple at first glance that it is hard to imagine them as the initiator in the virus—host cell interaction. Yet viruses have the capacity to manipulate the cell they are infecting into responding in ways that move the virus through its replication cycle. Often the action of the virus is subtle, consisting of tight binding of viral mole- cules to host cell molecules or production of an enzyme that alters the bal- ance between host cell and viral replication processes. This is evident from the very beginning of the influenza virus replication cycle. The virus first attaches to its target mammalian cell by means of a protein called hemag- glutinin, one of its envelope proteins. This protein was named for its ability Viruses of Mammalian Cells 20! 202 Chapter 10 A Nucleoprotein core Viral envelope protein Viral envelope and hon cell membrane fused -9'-. ..... .. Host cell membrane ‘—v—’ ‘—v—’ Viral envelope protein Viral envelope and Nucleoprotein core released binds host cell receptor host cell membrane into host cell cytoplasm begin to fuse Virus binds receptor on host cell: stimulates cell to take up virus by endocytosis Vesicle becomes acidified: matrix proteins destabilized Viral envelope fuses to vesicle membrane Nucleoprotein core released into cytoplasm Figure IOJ Entry and uncoating of viruses. The viruses in this figure have an envelope, but similar processes occur in the case of naked viruses. (A) Some viruses attach to the host cell receptor molecule they recognize, then fuse with the host cell cytoplasmic membrane and introduce their nucleoprotein core into the cytoplasm of the host cell. (B) Influenza virus and a number of other viruses use an internalization process in which the virus triggers its own uptake in an endocytic vesicle. After fusion of the viral surface with the vesicle membrane, the nucleoprotein core is released into the cytoplasm. to agglutinate red blood cells. Hemagglutinin proteins form trimers that project out from the viral surface (Fig. 10.4), making the viral surface appear studded with spikes. These glycoprotein spikes bind very specifically to sialic residues on the surface of a mammalian cell. Sialic acid is a sugar found widely on mammalian cells (Fig. 10.5). Because many mammalian cells have sialic acid residues on their surfaces, influenza viruses can infect many different cell types. Other viruses that have surface proteins which recognize molecules found only on the surfaces of a limited number of cell types have a more restricted range of cells they can infect. Thus, for exam- ple, HIV infects only certain cells of the immune system, because HIV sur- face proteins recognize molecules found on the surfaces of these particular cells but not on other types of cells. Another level of specificity is the ability of the virus to replicate once it enters the cell. Some cells are permissive for replication of a particular virus but not for replication of others. The model for binding of a viral surface protein to a molecule on the host cell (viral receptor) usually depicts a single viral protein interacting with a single receptor molecule on the host cell. Recently, in the case of HIV, it has become clear that some viruses may have more complex interactions with host cell surfaces. HIV surface proteins have a primary receptor, a pro- tein called CD4, which is found on certain cells of the immune system, but, before the invasion process can proceed, stabilization of the interaction between the viral surface protein and CD4 requires interaction between HIV and secondary receptor proteins called coreceptors (Chapter 18). It is now evident that the use of both a receptor and a coreceptor may be much more common among viruses than was thought previously to be the case. In the case of influenza virus, the simpler model of the attachment process, in which a single viral protein binds to a single mammalian cell molecule, seems to be an accurate representation of the interaction. Binding of the viral envelope protein spike to sialic acid is very tight and is stable enough to trig- ger the mammalian cell to initiate endocytosis in the vicinity of the bound virus, forming an endocytic vesicle that contains the virus {Figure 10.313). Endocytosis is a normal pathway used by mammalian cells to ingest nutrients. The virus in effect subverts this normal pathway to facilitate Figure l0.4 Envelope proteins of influenza virus. One of the envelope proteins {hemagglutinin} is involved in attachment of influenza virus to sialic acid residues on the host cell surface. Three hemagglutinin proteins are organized to form a pro- tein spike, which actually mediates attachment to sialic acid residues. The second envelope protein (neuraminidase) cleaves the sialid acid residues from carbohy- drate residues on the cell surface, thus releasing the virus from the cell. Four neu- raminidase proteins form a unit in the envelope. Hemagglutinin Neuraminidase Phospholipid bilayer Matrix proteins Viruses of Mammalian Cells 203 Figure “1.5 Structure of sialic acid. Sialic acid is a sugar found on the sur— face of most human cells, where it is attached covalently to other sugars. H H—(IZ—OH H—CIZ—OH H—‘é—OH 0 II 0 H3C 'C_ N H COO— HH HOH OHH 204 Chapter 10 entry into the mammalian cell. After the endocytic vesicle is formed, it begins to acidify. Acidification leads to a conformational change in the matrix proteins that underlie the envelope, allowing the envelope to be removed and the nucleoprotein core to be released into the cell cytoplasm. As already mentioned, not all viruses enter cells via the endocytic path- way. Some attach to and fuse directly with the cytoplasmic membrane of the mammalian cell, releasing the nucleoprotein core into the cytoplasm. A subset of viruses can use such a fusogenic pathway to move directly from an infected cell to a new cell without being released into the external envi- ronment. Such viruses have the advantage that, as long as they are inside cells, they are hidden from the immune system. Replication of the viral genome and production of viral proteins. After release into the cytOplasm, the influenza viral core moves to the nucleus, where it gains access to the interior through pores in the nuclear membrane. Normally, proteins that transit nuclear membrane pores are restricted to those types of proteins that need to enter or leave the nucleus, but viral nucleoprotein core proteins are able to mimic these proteins sufficiently well to pass this cellular barrier. Once inside the nucleus, the viral genome begins to replicate. Most RNA viruses replicate in the cytoplasm, whereas DNA viruses replicate in the nucleus. Influenza virus is an exception to this rule. Influenza virus has a segmented genome made of single-stranded RNA seg- ments. The sequence of these segments is the complement of the messenger RNA (mRNA) strand that will move into the cytoplasm to be translated into viral proteins. The mRNA strand is called the +strand, and the complement is called the —strand. Because the influenza virus genome is composed of —strands of RNA, these strands must first be copied into +strands, which will then be translated to produce viral proteins. This task is accomplished by an RNA-dependent RNA polymerase (replicase) encoded by the viral genome. The subunits of the replicase are proteins of the nucleoprotein core. Replicase makes a +stranded copy of the —strand. Some of these +strands become mRNAs, which are translated in the cytoplasm, and some remain in the nucleus, where the replicase uses them as a template to make new copies of the —stranded viral genome segments. The replicase subunits are contained in the nucleoprotein core and are thus available from the start. In general, viruses with —strand RNA genomes must bring preformed RNA replicases with them to make +strands, which can be translated to provide more RNA polymerase subunits. A virus with a +strand RNA genome may be able to dispense with preformed replicase, because it can be translated directly, making its replicase on the Spot. DNA viruses can use host enzymes for replication and transcription, although some provide their own replicases. Replication strategies for viruses with +strand RNA genomes, double-stranded RNA genomes, and DNA genomes are summarized in Figure 10.6. Influenza virus must make some copies of its —strand genome that will serve as mRNA and some that will serve as templates for replicase to pro- duce more —stranded copies of the genome. The virus solves this problem by a unique method of subverting normal host function. Normal mam- malian cell mRN As have an unusual structure on their 5' ends, called the 5' cap (Fig. 10.7A). A 5' cap signals the mammalian cell translation machinery that this mRNA is targeted for translation. Instead of making such a cap structure for its own +strand, the influenza virus replicase, once it has bound to the —strand viral RNA, acts as an endonuclease to cleave the 5‘ cap Figure l0.6 Overview of viral replication A ssRNA viruses Strategies. (Al +slrand RNA viruses, which are fitmnd arenml RNA m RNA replicaSE. capsid already in the messenger RNA (mRNA) form, p and EnVElOPe Dwain-i can be translated to produce viral proteins, such as the replicase that will be used to mpro- duce the viral genome and make more viral mRNA for translation. (B) —strand RNA viruses require a preformed replicase, which was part of the nucleoprotein core, to make the mRNA copy of the -strand. Double-stranded RNA viruses use a replication strategy similar to that of the —stranded RNA viruses. (C) Retroviruses (such as HIV) make a DNA copy of their RNA genome using a special viral enzyme, reverse transcriptase, which is brought preformed into the infected cell as part of the nucleoprotein core. (B) DNA Viruses usually use host cell enzymes for replication. Replicase l ~strand Replicase l +strand progeny RNA Capsid and envelope proteins Progeny virions B ssRNA (—strand} or dsRNA viruses Preformed re Iicase in virus Translation wstrand parental RNA ——P—h- +strand —.- Capsid and envelope proteins Preformed replicase l —strand progeny RNA Capsid and envelope proteins Progeny virions C Retroviruses +strand parental RNA 9reformed reverse transcriptase l ssDNA copy Preformed reverse transcriptase l dsDNA 1 Host RNA polymerase Reverse tmnscriptase. capsid Integrate: into host genome fitmnd RNA acts as mRNA —b- and envelope proteins +strand progeny RNA Capsid and envelope proteins 1 Progeny virions D Most DNA viruses Parental dsDNA my. Viral polymerase (if host polymerase not used} Capsid and envelope proteins Host or viral DNA polyerrnase l Progeny dsDNA Capsid and envelope proteins 1 Progeny virions 205 mRNA 0 HN \ O O 0 H1“ N u II II —o—p—|lv—I|>—o— I 0-0- 0- O=iI’——O— H2 0 O- o—<:H3i cl) oze—o— H3 0 O- OH c? o=e—o O. B Replicase Viral RNA (—strand) 5’ cap Host mRNA Endonuclease cleavage removes 5’ cap Viral RNA (—strand) Figure IOJ’ Structure of the 5' cap and the mechanism by which influenza virus subverts the host cell's translation machinery to its own use. (A) Structure of the 5' cap that identifies Replicase copies —strand eukaryotic messenger RNAs (mRNAs) for translation. {B} The replicase of influenza virus binds to the viral —strand RNA, then attaches a 5' cap from a host mRNA. Endonucleolytic activity of the viral replicase clips off the 5' cap, releasing the rest of the host cell mRNA. The 5' cap then primes synthe- sis of the +strand by the viral reph'case, resulting in a capped viral message. This strategy is unique to influenza virus. ‘— Host mRNA Viral RNA (—strand) Capped viral RNA {+strand) {mRNA} 206 n————_fl Viruses of Mammalian Cells 20? structure plus 10—12 nucleotides from a host mRNA molecule (Fig. 10.73). The captured cap region then primes synthesis of the +strand by the repliw case, resulting in a capped +strand copy of the viral genome. This strategy accomplishes two goals. First, it targets certain viral +strands for transla- I tion. Second, it helps shut down translation of mammalian cell messages, thus leaving the translation machinery of the infected cell to devote itself to synthesizing viral proteins. This “theft” strategy is one way to divert the host cell biosynthetic machinery to focus on translating viral transcripts and reproducing viral genomes instead of replicating the host cell genome and synthesizing host cell proteins, and all viruses must have some way of doing this. Some take advantage of their small genomes to replicate so rapidly compared to the host cell genome that they swamp the nucleic acid replication machinery of the host cell. In a similar way, their transcripts soon dominate in number, - thus monopolizing the host cell translation machinery. Other viruses inter fete with proteins that control host cell DNA or RNA synthesis, making it very inefficient compared to the more efficient synthesis of viral messages and genomes. Whatever the strategy used by the virus to enhance its own replication at the expense of normal host cell functions, the usual effect is that host cell biosynthesis of its own nucleic acids and proteins declines drastically and synthesis of viral genome and viral proteins predominates. Such drastic effects on host cell biosynthetic functions can have an undesirable effect from the virus’s point of view: apoptosis, a type of cell death, can be trig- gered, and the eukaryotic cell shuts down all its functions and dies. Some viruses counter ap0ptosis to keep the eukaryotic cell viable, from the biosynthetic point of view, long enough to give the virus time to complete its replication cycle. I Assembly and departure of virus particles from the infected mam- maiian cell. As replication of viral genomes and translation of viral proteins proceeds, copies of viral genomic segments and viral proteins begin to accu- mulate in the cytoplasm of the infected cell. When concentrations of these components are sufficiently high, the proteins of the nucleoprotein core interact with viral genome segments. Most RNA viruses have single genome segments, not 7~8 as in the case of influenza virus. The way in Which influenza virus manages to gather and keep track of its 7—8 genome segments is not known. Sometimes the virus makes a mistake and packages a "wrong" segment. The significance of this mistake for influenza virus evo- lution and for human health is described in Chapter 16. For now, suffice it to say that this is the origin of the strains of influenza virus that have been called “killer influenza” because they are no longer recognized by the human immune system. Once the nucleoprotein core has been assembled, the next step is to acquire the matrix proteins and envelope. While viral particles have been assembling in the cytoplasm, the influenza virus envelope proteins, the hemagglutinin and the neuraminidase, have been inserted into the cell membrane. The viral particles begin to bud out of the cell through the cyto- plasmic membrane, acquiring their envelope in the process (Fig. 10.8). At this point, the virus faces one final problem. The hemagglutinin spikes that allowed the virus to bind so tightly to the mammalian cell surface in the initial steps of infection now become a potential burden, because, as the virus buds through the membrane, the hemagglutinin spikes can bind 208 Chapter 10 / Hemagglutinin m ( %— Neu raminidase “1,, f (9 v e RNA genome segments Matrix proteins + nucleocapsid proteins Hemagglutinin and neunminidase inserted in host cell membrane; matrix proteins attach to hemagglutinin and neuraminidase in the cell membrane. As virus buds out of host cell. the envelope Viral genome segments associate wid-I is formed from the cell membrane containing matrix protein attached to hemagglutinin hemagglutinin and neuraminidase. and neuraminidase in host cell membrane Figure “3.8 Budding of influenza virus through the host cell cytoplasmic mem- brane. Viral envelope proteins localize to the cytoplasmic membrane as they are made. Note that each genome segment has its own nucleoeapsid proteins bound to the RNA segment rather than a standard capsid. These particles interact with viral matrix proteins and the envelope proteins as the virus begins to exit the cell. The virus acquires an enve10pe, complete with vitally-produced envelope proteins, in the process of budding. once again to sialic acid residues on the mammalian cell surface, tethering the virus to the cell it is trying to leave. The other envelope protein, neu- raminidase, is thought to be the solution to this problem. N euraminidase is an enzyme that cleaves the bond attaching sialic acid residues to other carbo- hydrate residues on the cell surface. This cleavage reaction helps to release the virus from the cell, so that the virus is now free to infect a fresh cell and repeat its replication cycle. Antiviral Compounds A review of the steps in viral replication shows that, although there are not many suitable targets for antiviral drugs, some do exist (Fig. 10.9). Stopping attachment of the virus to the host cell is a very attractive drug strategy, because if the virus never attaches to and enters the cell it will not be able to cause any damage. One way to stop attachment would be to provide mole- | Viruses of Mammalian Cells 209 Attachment mRNA synthesis Q .: "II-rd I ’._._""..___,.—_."'_, Genome replication-E .1 Translation (syndlesls Figure 10.9 Targets for antiviral compounds. Xs mark the steps that are targeted by currently available viral compounds. Because viruses differ from each other in a variety of ways, an antiviral compound that works for one type usually does not work for other x -Targets of antivirai compounds types 0f Vin-13- cules that resemble the viral receptor on the host cell surface enough to fool the virus into binding the receptor mimic instead of the cell itself. In the case of influenza virus, attempts have been made to use free sialic acid to pre- vent viral attachment. The problem with this approach is that the receptor mimic must be provided continuously and must have access to tissues the viruses are likely to infect. A more successful strategy for inhibiting attach- ment, vaccines that elicit an immune response against viral surface pro- teins, will be described in Chapter 14. For viruses that are endocytosed and uncoat in endocytic vesicles, the uncoating process is a potential target. In fact, a successful anti-influenza drug, amantidine, inhibits uncoating of the virus. Amantidine does not interfere with the process of endocytosis——it would be toxic if it did—but instead binds to influenza virus matrix proteins and prevents the conforma- tional change that is essential for uncoating. For viruses with replication enzymes that are different from host cell enzymes, viral replication is a suitable drug target. Often the compounds used to inhibit viral replication enzymes are analogs of nucleotides, which prematurely terminate synthesis of viral genomes or viral messages or otherwise interfere with viral replication. Examples of such antiviral com- pounds and how they work are given in later chapters. A cornerstone of HIV therapy is a collection of such analogs that are preferentially incorpo- rated by the HIV enzyme reverse transcriptase. Another unusual feature of the HIV replication process that serves as a target for a different type of antiviral drug is the formation of large polyproteins, which must be clipped apart by viral proteases to become activated (Chapter 18). The protease inhibitors used to treat HIV patients target this activity, not seen in human cells. Currently, viral attachment, uncoating, and replication are the main targets of antiviral compounds. It is conceivable that as more is learned about the replication cycles of viruses, new targets will be found. |_____ ZIO Chapter 10 Pathology of Viral Disease Explaining symptoms One factor that dictates the symptoms a virus causes is the type of host cell it infects. This specificity explains why hepatitis virus only infects liver cells and HIV targets certain cells of the immune system. In the case of influenza virus, the virus binds a receptor found on virtually all mammalian cells. Thus, some other factors must determine specificity because, in humans, influenza is normally a disease of the respiratory tract (Chapter 16). Birds can shed avian influenza virus from their intestinal cells, so clearly influenza is capable of attacking cells other than those of the respiratory tract. Why, then, is influenza usually localized to the cells of the human upper airway? One reason is that in humans the spread of influenza virus is by airborne transmission or by trans- mission of virus from hands to face and thence to the respiratory tract. Opportunity to encounter susceptible host cells is clearly a factor here. A Second factor is probably the human immune response. Most people who encounter a strain of influenza virus have probably encountered a similar strain previously. Thus, the immune response kicks in early on, keeping the infection localized to the site of infection. It may not be completely effec- tive at first if the new strain is not identical to the previously encountered one, but it may work early enough to allow the body to stem the infection. The symptoms of ordinary influenza are probably caused by the body’s response to dying human cells (inflammation; Chapter 12). In an early response to infec- tion, the human body mobilizes proteins called cytokines, which organize the defense responses of the body, together with cells that kill infected cells. This complex response to infection is the cause of generalized flu symptoms such as chills, fever, joint and muscle aches, and malaise. Thus, symptoms are not the result of viruses infecting the brain or the joints but result from the sec- ondary effects of viral destruction of respiratory cells. The importance of a timely immune response in limiting the spread of influenza virus is underscored by what happens when the human body encounters a strain of influenza virus that is completely new to the immune system. The influenza epidemic of 1918 and 1919, which killed millions of people worldwide is an example. Since that time, other outbreaks of “killer flu” have occurred with some regularity, but none quite so devastating or involving so many people. The cause of death in infected people is still somewhat controversial, because scientists and physicians at that time did not have the advanced diagnostic and analytical technologies we have today. Many of the deaths were undoubtedly caused by secondary bacterial infections made possible by the weakened defenses of the airway (Chapter 16). Nonetheless, some evidence suggests that the “killer flu” strains pene- trated deeper into the lung than normal influenza infections and were thus more destructive. This could be explained by the delayed immune response that allowed the virus to gain access to susceptible cells it would not nor- mally have had a chance to encounter. Whatever the explanation for “killer flu," it is clear that the symptoms and severity of a viral disease depend on a constellation of factors. These include not only the specificity of a virus for a certain type of mammalian cell but also the route of transmission and the immune status of the person encountering the virus. Outcomes of a viral infection Symptoms of a viral infection are also influenced by the fact that there can be several possible outcomes of an initial infection event (Fig. 10.10). After Virus contacts host cell l—l—l Nonproductive infection (no progeny viruses produced) l—;| Viral genome persists in host cell Productive infection (progeny viruses produced) l—l—l Cell dies.many Cell survives.sheds virions produced low number of virions (acute infection) (chronic infection) Cytopathic effects Latent Infection Vlral genome lost initial entry of a virus into the body, the viral infection may remain local- ized, either because only certain cell types are affected (e.g., liver cells, gas- trointestinal cells) or because the immune response quickly brings the infection under control. If the viral infection is cleared, symptoms will dis- appear. If viruses enter the bloodstream, the infection may spread through- out the body (systemic infection). Viruses can either spread as free viral particles in blood or by infecting the phagocytic cells that constantly move through the bloodstream (white cells; Chapter 12). These phagocytic cells are intended to be protective, but, if they become infected with viruses, they can actually help to spread the infection to distant parts of the body. This is true of HIV, which is capable of infecting a type of mobile phagocytic cell called the monocyte. Both localized and systemic infections can become latent. A latent infection differs from clearance of viruses in the body in that, although symptoms have abated, the virus is still present. In the latent form, the virus is temporarily inactive and has been cleared from cells where it can repli~ cate but is still capable of breaking out of the latent state to cause infection in the future. A good example of this latency is provided by herpes simplex virus, which causes lesions (cold sores, genital sores) when it is actively replicating in epithelial cells but has a latent stage in nearby neurons (Chapter 18). Emergence from this latent stage to reinfect epithelial cells prompts recurrences of herpes simplex symptoms. The outcome of an infection caused by a particular virus can vary sub- stantially from person to person (see Box 10—] for some episodes that illus- trate this dramatically). To some extent, this variation can be understood on the basis of the immune status of the person. A person who mounts a vigor» ous immune response is less likely to develop symptoms than a person who does not. Yet this cannot be the only explanation, because people with simiv lar levels of immune competence can still exhibit great variations in the severity of symptoms. The number of viruses to which the person is exposed and the route of exposure are certainly important. Symptoms resulting from exposure to a lower number of viruses are more easily brought under control than are those from exposure to a higher number. Underlying conditions, such as concurrent infection with another microbe or conditions like diabetes that can weaken the immune system, can play an important role. The immune response may actually contribute to disease Viruses of Mammalian Cells 2| | Figure 1010 Possible outcomes ofa viral infection. 2l 2 Chapter 10 hox_________— 10—1 Two Episodes of Unintentional Inoculation with Live Viruses I linstrate the Range of Susceptibility to Viral Infections Case I. The Cutter Incident. In the I9SOs.before the development of a live polio vaccine.the polio vaccine consisted of formalin-killed viruses (Salk vaccine).A shipment of polio vaccine prepared by Cutter Laboratories in I955 was not properly treated and contained live polio virus. Before this was discovered.nearly l20.000 schoolchildren were injected with the contaminated vaccine. Estimates based on a subsequent follow~up investi- gation and on information about the inci- dence of natural immunity in the US. population indicated that about half of the children inoculated with the vaccine already produced antibodies against the polio virus and were protected from con- tracting the disease. Approximately 25% of the remaining children were infected. as indicated by the appearance of symp- toms or shedding of the virus in feces. but only 60 cases of paralytic polio (the most severe form of the disease) actually developed. Case 2. Yellow fever episode. During World War ll, yellow fever was a serious problem for US. troops sent to coun- tries where the disease was endemic. Most military personnel were vaccinated to prevent yellow fever. In 1942, a ship— ment of vaccine that had been uninten- tionally contaminated with hepatitis B virus was administered to 40 l .535 mili- tary personnel. Of these. only 9 l4 {about 0.2%) developed jaundice.and fewer than 50 developed the severe form of hepatitis. When considering the figures quoted above. it should be kept In mind that the cases was low. Exposure to higher levels of virus,as might happen during an epi- demic, would result in a higher incidence of disease. What these cases illustrate. however. is the broad range of Suscepti- bility even in populations that are rela- tively homogeneous wid-I respectto age and standard of Ilving. Sources: Nathanson. N.. and A. Langmuir. I963.The Cutter incident Poliomyelltis following formaldehyde- inactivated poliovirus vaccination in the United States during the spring of I955. Amen]. Hyg. 78: I 6-8 I. Sawyer.W. at al. i944.]aundice in army personnel in the western region of the United States and its relation to vaccina~ tion against yellow fevecAmerJ. Hyg. 39: I, amount of virus actually injected in these 337—430. severity if the immune response mounted against an initial infection is inappropriate and exacerbates symptoms the second time the person is exposed. This seems to be the case with dengue virus infections, in which the second exposure produces worse disease than the first (Chapter 20}. Such aberrant immune responses are uncommon. but they illustrate the complexity of the range of factors and conditions that mix together to deter- mine the severity of symptoms. Disease severity also may have a genetic component. For example, peo- ple with genetic defects in the gene encoding the coreceptor needed by HIV for effective attachment to human cells take longer to develop symptoms or may not develop symptoms at all (Chapter 18). Is this an HIV~specific phe- nomenon or are there genetic factors that affect the outcome of other viral infections? We need a better understanding of person-to-person variations in the severity of symptoms of viral diseases. Why Aren’t We Doing Better? It is perhaps understandable that our attempts to understand disease- causing bacteria and eukaryotic microbes have foundered on the realization that these are formidable enemies—not only because of their adaptability but because they are so complex, both genetically and metabolically. Failures in this area are easy to rationalize. Yet, how can we rationalize simi- lar failures to deal with microorganisms so simple that even the most primi- tive genome sequencing facility could sequence their genomes completely in a couple of weeks at the most? The answer is that their apparent simplic— ity is highly misleading. As isolated entities, these microorganisms are sim- ple in the sense that they are composed, in most cases, of a very short shop— ping list of genes and proteins. What allows this apparent simplicity to balloon into complexity as great as that of the cells the viruses infect is the ability of viruses to take advantage of characteristics of eukaryotic cells in Ways that can produce very complex responses. To revert to the words of Jonathan Swift at the beginning of the chapter, just as a flea biting a person can touch off all sorts of responses, from mental feelings of itching and irri- ta tion to results of infectious agents injected by the bite, something as sim- ple as a virus has only to tweak a mammalian cell in the right way to elicit a cascade of responses, which, if understood, would lead to a far deeper com- prehension of both viral and mammalian cell function. Viruses of Mammalian Cells 2|3 320 Chapter 16 Influenza, Still a Serious Health Threat Importance of influenza Influenza is estimated to cause about 20,000 deaths per year in the United States alone. Generally, death is caused not by the influenza virus itself, but by the subsequent bacterial pneumonia that develops in some people after a bout of influenza. In most years, the vast majority of fatalities occur in elderly people, but there have been influenza epidemics, such as that of 1918, when millions of people of all ages died. Subsequently, there have been two or three other exceptionally bad flu years. The last outbreak of “killer flu” occurred in the late 19705. Public health officials are concerned about when the next epidemic will strike and are planning for coping with it when—not if—it comes. One important advance has been the develop- ment of a fairly effective vaccine against influenza, a vaccine that public health officials hope will take the punch out of the next wave of killer flu. As explained in Chapter 14, however, the Achilles heel of this vaccine is the continued and unpredictable evolution of new strains of the virus that will arise due to its high rate of mutation. This, together with the months-long lag time between detection of a new strain of influenza virus and delivery of a vaccine to the public, makes the vaccine less of a protection against a future flu epidemic than it could be. Influenza is caused by an enveloped virus, the influenza virus. There are three types of influenza virus: A, B, and C. They differ mainly in the amino acid sequences of their envelope proteins. Type A is most likely to cause the large, multination outbreaks called pandemics. Types B and C cause milder disease and usually do not produce epidemics. The structure of influenza virus and its replication cycle were introduced in Chapter 10. To recap briefly, the envelope of influenza virus contains two types of pro- teins, neuraminidase and hemagglutinin. Hemagglutinin allows the virus to enter human cells by attaching to their sialic acid residues. Neu- raminidase is important for viral exit from the cell. Both hemagglutinin and neuraminidase are the target of anti-influenza vaccines. The influenza genome has an interesting and significant property: it is not a single seg- ment but is composed of 7—8 segments of single-stranded RNA. Each of these segments has its own protein covering. This segmented form of the genome makes possible large genetic changes in the virus, a phenomenon called antigenic shift. Influenza viruses attack the ciliated epithelial cells of the respiratory tract. As already mentioned, these cells are an important defense against infection of the lung, and impairment of this defense increases the likeli- hood that bacteria can reach the lung and cause a secondary infection. Antigenic drift and antigenic shift Influenza virus is constantly changing. In fact, it is the fastest mutating virus known, with another RNA virus, HIV, as a close second. Viruses with RNA genomes generally mutate faster than viruses with DNA genomes, because the enzymes that reproduce RNA are less accurate than the ones that reproduce DNA. Small changes in the viral genome sequence can lead to changes in the envelope proteins, making them different enough to be less well recognized by the memory cells of the immune system (antigenic drift). People are advised to get flu vaccine each year, not just to bolster the immune response, but because the composition of the flu vaccine is The Lung, aVital but Vulnerable Organ 32I changed from time to time to reflect differences in the surface proteins of the viruses currently circulating. Small changes resulting from mutations do not eliminate completely the protection conferred by an old vaccination or by infection with previous years’ influenza strains. Thus, more people may get the flu when a new variant of influenza virus arrives that is only slightly different from the previous strain, but the disease should not be more severe, because of the partial protection from the earlier exposure. “Killer flu" arises from big genetic changes that make the virus virtually invisible to the immune system, so that no protection remains (antigenic shift). Big changes in envelope proteins occur because of two features of the influenza virus: its segmented genome and the ability of influenza A viruses to infect many kinds of mammals and birds. Suppose, for example, that a pig is infected simultaneously with human influenza A virus and a porcine influenza virus. If, during the co-infection, the two types of virus replicate in the same cell, the predominantly human virus can pick up a genome segment from the avian virus (Fig. 16.2). This may result in a new influenza A virus with new envelope proteins that will not be recognized by the immune systems of most people in the population. The "new" virus can cause the most serious form of influenza. Scientists using modern molecu- lar techniques have amplified and cloned a DNA copy of influenza genome segments from tissue specimens taken during the 1918 pandemic. By com- paring the nucleotide sequence of the cloned DNA copies of these segments with sequences of human and animal influenza viruses, they concluded that the nucleic acid sequences of some of the 1918 strain genome segments matched pig influenza genome sequences more closely than human influenza genome sequences. This suggests that the 1918 virus contained a genome segment of pig origin. Mixing of viral genome segments occurs in other animals, too, such as the ill-fated Hong Kong chickens. Bacterial Pneumonia Importance of bacterial pneumonia S. pneumoniae, also called the pneumococcus, is the most common cause of bacterial pneumonia and is frequently the cause of death (as a result of sec- ondary infection) in fatal influenza cases. S. pneumoniae has an impressive record. According to the CDC, each year in the United States alone this one species of bacterium causes 3,000 cases of meningitis, 50,000 cases of bloodstream infection, 500,000 cases of bacterial pneumonia, and 7 million cases of ear infection. Not bad for a tiny bacterium, which, when viewed under a microsc0pe, looks about as threatening as a beach ball. S. pneumo— niae is a gram-positive coccus often seen in pairs (diplococcus) (Fig. 16.3). lts distinctive diplococcal appearance is useful for rapid diagnosis of the disease. Two other bacteria that cause pneumonia are Klebsiella pneumoniae and Haemophilus influenzae (so named because it was originally misidenti- fied as the cause of influenza). Both of these bacteria are gram-negative rods. Gram-orientation notwithstanding, all three of these bacteria cause the same type of lung damage and symptoms, although Klebsiefla pneu- monia is somewhat more destructive than the other two. Why, then, would a physician care which one of these bacteria was causing the disease? Until recently, S. pneumoniae was a lot less likely to be resistant to antibiotics than K. pneumoniae and H. influenzae, and good old 322 Chapter 16 A Virus | Virus 2 human influenza virus from another species of animal B Virus 1 I Assembly of new Virus 2 virus particles New viruses released from cell New variant virus against which there is no protection Same as original virus 2 Same as original virus I Figure l6.2 Antigenic shift in influenza virus. (A) Influenza viruses from differ- ent species simultaneously infect a single animal. (B) The two types of viruses repli- cate in a single cell. When the new viral particles are assembled, segments of the genome from different viruses occasionally are incorporated into the same virus particle. If a human influenza virus receives a genome segment from a swine or avian virus, this “new” virus may have new envelope proteins that allow it to bypass the immune system of mOst of the human pepulation. penicillin was usually effective. By contrast, more advanced drugs were needed to treat the gram-negative species. In recent years, the appearance of penicillin-resistant S. pneumoniae has changed this picture somewhat, but at least in certain regions of the United States, penicillin still works for ...
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This note was uploaded on 04/29/2008 for the course CHEM 227 taught by Professor Santander during the Spring '08 term at Texas A&M.

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