简介
Designed for students learning about viruses for the first time(t the undergraduate or graduate level), Fundamentals of Molecular Virology is presented in a style which relates to today′s students and professors. The text approaches learning about virology by presenting a set of chapters each of which covers a specific virus family, using one or two well–studied viruses as examples. Each chapter is designed to tell a story about the virus under discussion, and to portray the "personality" of that virus. The text incorporates lessons from classic and contemporary concepts providing a well–rounded presentation on the subject of virology.
FEATURES OF FUNDAMENTALS OF MOLECULAR VIROLOGY
? Unique, Applied Chapter Stories. Each chapter presents a unique example or case to help introduce the students to the different viruses that will be studied or examined in that chapter.
? Evolutionary Boxes. Feature exciting and current developments in molecular virology. These are integrated throughout the entire book and can be found in every chapter. These help students understand the importance of currency and application of virology.
? Comprehensive, Illustrative Art Program. The text contains a number of two–color figures which focus on the individual steps in virus replication and helps draw student′s attention to important concepts and details.
? Coverage of Human Pathogens. Includes chapters that cover important human pathogens such as smallpox virus, measles virus, poliovirus, herpes viruses, human immunodeficiency virus, hepatitis B virus, Ebola virus, SARS corona virus, West Nile virus, and others.
What instructors are saying about Fundamentals of Molecular Virology
"I like the structured treatment that is presented in Acheson. Overall, it is one of the best written and clearly organized texts on the subject I have seen." – Jeannine Williams, College of Marin
"I found the text very readable and believe it will appeal to a wide audience of students...I believe this text will have broad appeal in a field where few texts exist." – Michael Roner, The University of Texas at Arlington
"The main strength of the book is the great molecular detail the author achieves, but still at a level that an undergraduate student should be able to master. I like the blend of molecular with medical; this has been lacking in most virology books that I have considered using." – Darlene Walro, Walsh University
目录
I. INTRODUCTION
1. Introduction to Virology 1
THE NATURE OF VIRUSES 2
Viruses consist of a nucleic acid genome packaged in a
protein coat 2
Viruses are dependent on living cells for their replication 2
Virus particles break down and release their genomes inside
the cell 2
Virus genomes are either RNA or DNA, but not both 2
WHY STUDY VIRUSES? 3
Viruses are important disease-causing agents 3
Viruses can infect all forms of life 3
Viruses are the most abundant form of life on Earth 4
The study of viruses has led to numerous discoveries in
molecular and cell biology 4
A BRIEF HISTORY OF VIROLOGY: THE STUDY OF
VIRUSES 5
The scientific study of viruses is very recent 5
Viruses were first distinguished from other microorganisms
by filtration 5
The crystallization of tobacco mosaic virus challenged
conventional notions about genes and the nature of living
organisms 5
The ?phage group? stimulated studies of bacteriophages and
helped found the field of molecular biology 7
Study of tumor viruses led to discoveries in molecular
biology and understanding of the nature of cancer 7
DETECTION AND TITRATION OF VIRUSES 8
Most viruses were first detected and studied by infection of
intact organisms 8
The plaque assay arose from work with bacteriophages 8
Eukaryotic cells cultured in vitro have been adapted for
plaque assays 8
Hemagglutination is a convenient and rapid assay for many
viruses 9
Virus particles can be seen and counted by electron
microscopy 10
The ratio of physical virus particles to infectious particles can
be much greater than 1 10
THE VIRUS REPLICATION CYCLE:
AN OVERVIEW 10
The single-cycle virus replication experiment 10
An example of a virus replication cycle: mouse
polyomavirus 11
Analysis of viral macromolecules reveals the detailed
pathways of virus replication 12
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STEPS IN THE VIRUS REPLICATION CYCLE 12
1. Virions bind to receptors on the cell surface 12
2. The virion (or the viral genome) enters the cell 12
3. Early viral genes are expressed: the Baltimore classification
of viruses 13
The six groups in the Baltimore classification system 13
4. Early viral proteins direct replication of viral genomes 14
5. Late messenger RNAs are made from newly-replicated
genomes 15
6. Late viral proteins package viral genomes and assemble
virions 15
7. Progeny virions are released from the host cell 15
2. Virus Structure 17
The molecular structure of virus particles 17
How virus structure is studied: viruses come in a variety of
sizes and shapes 18
Small viruses come in simple, symmetrical packages 18
Many virus capsids have icosahedral symmetry 18
Some examples of virions with icosahedral symmetry 21
The concept of quasi equivalence 21
How many subunits can be accomodated on the capsid
surface? 22
Other structures, large and small, display icosahedral
symmetry 23
Many virus capsids are organized as helical tubes 23
Larger viruses come in more complex packages 24
Specific packaging signals direct incorporation of viral
genomes into virions 25
Core proteins may accompany the viral genome inside the
capsid 25
Scaffolding proteins help in virion assembly but are not
incorporated into the mature virion 25
Viral envelopes are made from lipid bilayer membranes 26
Viral glycoproteins are inserted into the lipid membrane to
form the envelope 26
Budding is driven by interactions between viral proteins 27
Assembly and disassembly of virions: the importance of an
irreversible step 27
3. Virus Classification: The World of
Viruses 30
VIRUS CLASSIFICATION 30
Many viruses, infecting virtually all known life forms, have
been discovered 30
Virus classification is based on molecular architecture,
genetic relatedness, and host organism 31
Contents
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Viruses are grouped into species, genera, and families 31
Distinct naming conventions and classification schemes have
developed in different domains of virology 32
MAJOR VIRUS GROUPS 32
Study of the major groups of viruses leads to understanding
of shared characteristics and replication pathways 32
Viruses with single-stranded DNA genomes are small and
have few genes 33
Viruses with double-stranded DNA genomes include the
largest known viruses 34
Most plant viruses and many viruses of vertebrates have
positive-strand RNA genomes 35
All viruses with negative-strand RNA genomes have helical
nucleocapsids and some have fragmented genomes 36
Viruses with double-stranded RNA genomes have
fragmented genomes packaged in capsids with icosahedral
symmetry 37
Viruses with a reverse transcription step in their replication
cycle can have either RNA or DNA genomes 38
Satellite viruses and satellite nucleic acids require a helper
virus to replicate 39
Viroids do not code for proteins, but replicate independently
of other viruses 39
THE EVOLUTIONARY ORIGIN OF VIRUSES 40
The first steps in the development of life on earth: the RNA
world 40
Viroids and RNA viruses may have originated in the RNA
world 40
The transition to the DNA-based world 40
Small and medium-sized DNA viruses could have arisen
as independently-replicating genetic elements
in cells 41
Large DNA viruses could have evolved from cells that
became obligatory intracellular parasites 41
4. Virus Entry 43
How do virions get into eukaryotic cells? 43
Enveloped and nonenveloped viruses have distinct
penetration strategies 44
Some viruses can pass directly from cell to cell 44
A variety of cell surface proteins can serve as specific virus
receptors 45
Receptors interact with viral glycoproteins, surface
protrusions, or ?canyons? in the surface of the virion 45
Many viruses enter the cell via receptor-mediated
endocytosis 45
Passage from endosomes to the cytosol is often triggered by
low pH 47
Membrane fusion is mediated by specific viral ?fusion
proteins? 47
Fusion proteins undergo major conformational changes that
lead to membrane fusion 48
Nonenveloped viruses penetrate by membrane lysis or pore
formation 49
Virions and capsids are transported within the cell in vesicles
or on microtubules 49
Import of viral genomes into the nucleus 50
The many ways in which viral genomes are uncoated and
released 50
5. Single-Stranded RNA Bacteriophages 53
The discovery of RNA phages stimulated research into
messenger RNA function and RNA replication 53
RNA phages are among the simplest known organisms 54
Two genera of RNA phages have subtle differences 54
RNA phages bind to the F-pilus and use it to insert their
RNA into the cell 55
Phage RNA is translated and replicated in a regulated
fashion 55
RNA secondary structure controls translation of lysis and
replicase genes 56
Ribosomes translating the coat gene disrupt secondary
structure, allowing replicase translation 57
Ribosomes terminating coat translation can reinitiate at the
lysis gene start site 57
Replication versus translation: competition for the same
RNA template 58
Genome replication requires four host cell proteins plus the
replicase 58
A host ribosomal protein directs polymerase to the coat start
site 59
Polymerase skips the first A residue but adds a terminal A to
the minus strand copy 59
Synthesis of plus strands is less complex and more efficient
than that of minus strands 59
The start site for synthesis of maturation protein is normally
inaccessible to ribosomes 61
Synthesis of maturation protein is controlled by delayed
RNA folding 61
Assembly and release of virions 62
6 Bacteriophage _X 174 63
_X174: a tiny virus with a big impact 63
Overlapping reading frames allow efficient use of a small
genome 64
_X174 binds to glucose residues in lipopolysaccharide on the
cell surface 65
_X174 delivers its genome into the cell through spikes on the
capsid surface 66
Stage I DNA replication generates double-stranded
replicative form DNA 66
Gene expression is controlled by the strength of promoters
and transcriptional terminators 66
Replicative form DNAs are amplified via a rolling circle
mechanism 67
Summary of viral DNA replication mechanisms 67
Procapsids are assembled by the use of scaffolding proteins
67
Contents xv
Scaffolding proteins have a flexible structure 68
Single-stranded genomes are packaged into procapsids as
they are synthesized 68
Role of the J protein in DNA packaging 69
Cell lysis caused by E protein leads to release of phage 69
Did all icosahedral ssDNA virus families evolve from a
common ancestor? 69
7. Bacteriophage T7 71
T7: a model phage for DNA replication, transcription, and
RNA processing 71
T7 genes are organized into three groups based on
transcription and gene function 72
Entry of T7 DNA into the cytoplasm is powered by
transcription 73
Transcription of class II and III genes requires a novel
T7-coded RNA polymerase 73
Class II genes code for enzymes involved in T7 DNA
replication 74
T7 RNAs are cleaved by host cell ribonuclease III to smaller,
stable mRNAs 74
Regulation of class III gene expression 74
DNA replication starts at a unique internal origin and is
primed by T7 RNA polymerase 75
Large DNA concatemers are formed during replication 76
Concatemers are processed and packaged into preformed
proheads 76
Special features of the T7 family of phages 76
8. Bacteriophage Lambda: A Piñata of
Paradigms 79
In the beginning . . . 80
Uptake of _DNA depends on cellular proteins involved in
sugar transport 80
The _lytic transcription program is controlled by
termination and antitermination of RNA synthesis at
specific sites on the genome 81
The CI repressor blocks expression of the lytic program by
regulating three nearby promoters: PL, PR, and PRM 82
Cleavage of CI repressor in cells with damaged DNA leads to
prophage induction 83
The Cro repressor suppresses CI synthesis and regulates
early gene transcription 83
Making the decision: Go lytic or lysogenize? 83
A quick review 85
Breaking and entering: The insertion of _DNA into the
bacterial chromosome 85
The great escape: The liberation of _DNA from the
bacterial chromosome 86
Int synthesis is controlled by retroregulation 86
_DNA Replication is directed by O and P, but carried out
by host cell proteins 87
Assembly of _heads involves chaperone and scaffolding
proteins 87
DNA is inserted into preformed proheads by an ATPdependent
mechanism 87
Host cell lysis 88
9. Parvoviruses 89
Parvoviruses have very small virions and a linear, singlestranded
DNA genome 89
Parvoviruses replicate in cells that are going through the cell
cycle 90
Discovery of mammalian parvoviruses 90
Parvoviruses have one of the simplest known virion
structures 91
Parvoviruses have very few genes 91
Single-stranded parvovirus DNAs have unusual terminal
structures 92
Uncoating of parvovirus virions takes place in the nucleus
and is cell-specific 92
DNA replication begins by extension of the 3_ end of the
terminal hairpin 93
The DNA ?end replication? problem 93
Steps in DNA replication 93
Non-structural proteins are multifunctional 95
Adenovirus functions that help AAV replication 96
In the absence of helper virus, AAV DNA can integrate into
the cell genome 96
Parvovirus pathogenesis: the example of B19 virus 96
10. Polyomaviruses 98
Mouse polyomavirus was discovered as a tumor-producing
infectious agent 98
Simian virus 40 was found as a contaminant of Salk
poliovirus vaccine 99
Polyomaviruses are models for studying DNA virus
replication and tumorigenesis 99
Polyomavirus capsids are constructed from pentamers of the
major capsid protein 99
The circular DNA genome is packaged with cellular
histones 100
Circular DNA becomes supercoiled upon removal of
histones 100
Supercoiled DNA can be separated from relaxed or linear
DNA molecules 101
Polyomavirus genes are organized in two divergent
transcription units 102
Virions enter cells in caveolae and are transported to the
nucleus 102
The viral minichromosome is transcribed by cellular RNA
polymerase II 103
Four early mRNAs are made by differential splicing of a
common transcript 104
T antigens share common N-terminal sequences but have
different C-terminal sequences 105
T antigens bring resting cells into the DNA synthesis (S)
phase of the cell cycle 105
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Small T antigen inhibits protein phosphatase 2A and induces
cell cycling 105
Middle T antigen stimulates protein tyrosine kinases that
signal cell proliferation and division 105
Large T antigen activates or suppresses transcription of
cellular genes by binding to a number of important cellular
regulatory proteins 107
Large T antigen hexamers bind to the origin of DNA
replication and locally unwind the two DNA strands 108
Large T antigen hexamers assemble cellular DNA synthesis
machinery to initiate viral DNA replication 110
High levels of late transcripts are made after DNA
replication begins 111
Three late mRNAs are made by alternative splicing 112
How do polyomaviruses transform cells in vitro and cause
tumors in vivo? 112
11. Papillomaviruses 114
Papillomaviruses cause warts and other skin and mucosal
lesions 114
Oncogenic human papillomaviruses are a major cause of
genital tract cancers 115
Papillomaviruses are not easily grown in cell culture 115
Papillomavirus genomes are circular, double-stranded
DNA 116
The infectious cycle follows differentiation of epithelial
cells 116
Viral mRNAs are made from two promoters and two
polyadenylation signals 117
Viral E1 and E2 proteins bind to the replication origin and
direct initiation of DNA replication 118
Viral E7 protein interacts with cell cycle regulatory proteins,
particularly Rb 118
Viral E6 protein controls the level of cellular p53
protein 120
Synergism between E6 and E7 and the predisposition to
cancer 121
Cells transformed by papillomaviruses express E6 and E7
gene products from integrated viral DNA 121
Future prospects for diagnosis and treatment of diseases
caused by papillomaviruses 121
12. Adenoviruses 123
Adenoviruses cause respiratory and enteric infections in
humans 124
Adenoviruses can be oncogenic, but not in humans 124
Virions have icosahedral symmetry and are studded with
knobbed fibers 124
Fibers make contact with cellular receptor proteins to initiate
infection 124
Expression of adenovirus genes is controlled at the level of
transcription 126
E1A proteins are the kingpins of the adenovirus growth
cycle 127
E1A proteins bind to the retinoblastoma protein and activate
E2F, a cellular transcription factor 127
E1A proteins also activate other cellular transcription
factors 128
E1A proteins indirectly induce apoptosis by activation of
cellular p53 protein 128
E1B proteins suppress E1A-induced apoptosis, allowing viral
replication to proceed 128
The preterminal protein primes DNA synthesis carried out
by viral DNA polymerase 129
Single-stranded DNA is circularized via the inverted
terminal repeat 130
The major late promoter is activated after DNA replication
begins 131
Five different poly(A) sites and alternative splicing generate
multiple late mRNAs 131
The tripartite leader ensures efficient transport of late
mRNAs to the cytoplasm 132
The tripartite leader directs efficient translation of late
adenovirus proteins 132
Adenoviruses kill cells by apoptosis, aiding virus release 132
Cell transformation and oncogenesis by human
adenoviruses 132
13. Herpes Simplex Virus 134
Herpesviruses are important human pathogens 135
Most herpesviruses can establish latent infections 135
Herpes simplex virus genomes contain both unique and
repeated sequence elements 135
Nomenclature of herpes simplex virus genes and
proteins 137
The icosahedral capsid is enclosed in an envelope along with
tegument proteins 137
Entry by fusion is mediated by envelope glycoproteins
and may occur at the plasma membrane or in
endosomes 138
Viral genes are sequentially expressed during the replication
cycle 138
Tegument proteins interact with cellular machinery to
activate viral gene expression and to degrade cellular
messenger RNAs 139
Immediate early ( _) genes regulate expression of other
herpesvirus genes 140
_gene products set the stage for viral DNA replication 140
Herpesvirus begins with bidirectional DNA replication 141
Rolling-circle replication subsequently produces multimeric
concatemers of viral DNA 141
DNA replication leads to activation of _1 and _2 genes 142
Viral nucleocapsids are assembled on a scaffold in the
nucleus 143
Envelopment and egress: three possible routes 143
Many viral genes are involved in blocking host responses to
infection 143
The establishment and maintenance of virus latency 145
Latency-associated transcripts include stable introns 145
Contents xvii
14. Baculoviruses 147
Insect viruses were first discovered as pathogens of
silkworms 148
Baculoviruses are used for pest control and to express
eukaryotic proteins 148
Baculoviruses produce two kinds of particles: ?budded? and
?occlusion-derived? virions 149
Baculoviruses have large, circular DNA genomes and encode
many proteins 150
Insects are infected by ingesting occlusion bodies; infection
spreads within the insect via budded virions 151
Viral proteins are expressed in a timed cascade regulated at
the transcription level 152
Immediate early gene products control expression of early
genes 152
Early gene products regulate DNA replication, late
transcription, and apoptosis 153
Late genes are transcribed by a novel virus-coded RNA
polymerase 153
Baculoviruses are widely used to express foreign
proteins 156
15. Poxviruses 157
Smallpox was a debilitating and fatal worldwide disease 158
Variolation led to vaccination, which has eradicated smallpox
worldwide 158
Poxviruses remain a subject of intense research interest 159
Linear vaccinia virus genomes have covalently sealed hairpin
ends and lack introns 159
Two forms of vaccinia virions have different roles in
spreading infection 160
Poxviruses replicate in the cytoplasm 162
Poxvirus genes are expressed in a regulated transcriptional
cascade controlled by viral transcription factors 162
Virus-coded enzymes packaged in the core carry out early
RNA synthesis and processing 162
Enzymes that direct DNA replication are encoded by early
mRNAs 163
Poxviruses produce large concatemeric DNA molecules that
are resolved into monomers 164
Postreplicative mRNAs have 5_ end poly(A) extensions and 3_
end heterogeneity 164
Mature virions are formed within virus ?factories? 165
Extracellular virions are extruded through the plasma
membrane by actin tails 166
Poxviruses make several proteins that target host immune
defenses 167
16. Picornaviruses 169
Picornaviruses cause a variety of human and animal diseases
including poliomyelitis and the common cold 170
Poliovirus: a model picornavirus for vaccine development
and studies of replication 170
Picornavirus virions bind to cellular receptors via depressions
or loop regions on their surface 171
Genome RNA may pass through pores formed in cell
membranes by capsid proteins 171
Translation initiates on picornavirus RNAs by a novel
internal ribosome entry mechanism 172
Essential features of picornavirus IRES elements 173
Interaction of picornavirus IRES elements with host cell
proteins 175
Picornavirus proteins are made as a single precursor polyprotein
that is autocatalytically cleaved by viral proteinases 176
Picornaviruses make a variety of proteinases that cleave the
polyprotein and some cellular proteins 176
Replication of picornavirus RNAs is initiated in a
multiprotein complex bound to proliferated cellular
vesicles 176
RNA synthesis is primed by VPg covalently bound to uridine
residues 177
Virion assembly involves cleavage of VP0 to VP2 plus
VP4 178
Inhibition of host cell macromolecular functions 179
17. Flaviviruses 181
Flaviviruses cause several important human diseases 182
Yellow fever is a devastating human disease transmitted by
mosquitoes 182
A live, attenuated yellow fever virus vaccine is available and
widely used 183
Hepatitis C virus: a recently discovered member of the
Flaviviridae 183
The flavivirus virion contains an icosahedral nucleocapsid
wrapped in a tightly fitted envelope 183
Flavivirus E protein directs both binding to receptors and
membrane fusion 184
Flaviviruses enter the cell by pH-dependent fusion 185
Flavivirus genome organization resembles that of
picornaviruses 185
The polyprotein is processed by both viral and cellular
proteinases 186
Nonstructural proteins organize protein processing, viral
RNA replication, and capping 187
Flavivirus RNA synthesis is carried out on membranes in
the cytoplasm 188
Virus assembly also takes place at intracellular
membranes 189
18. Togaviruses 191
Most togaviruses are arthropod borne, transmitted between
vertebrate hosts by mosquitoes 192
Togavirus virions contain a nucleocapsid with icosahedral
symmetry wrapped in an envelope of the same
symmetry 192
Togaviruses enter cells by low pH-induced fusion inside
endosome vesicles 193
xviii Contents
Nonstructural proteins are made as a polyprotein that is
cleaved by a viral protease 193
Partly-cleaved nonstructural proteins catalyze synthesis of
full-length antigenome RNA 194
Replication and transcription: synthesis of genome and
subgenomic RNAs 196
Structural proteins are cleaved during translation and
directed to different cellular locations 196
Assembly of virions and egress at the plasma membrane 197
Effects of mutations in viral proteins on cytopathic
effects and on pathogenesis 198
Alphaviruses have been modified to serve as vectors for
the expression of heterologous proteins 199
Alphavirus vectors have multiple potential uses 199
19. CORONAVIRUSES 201
Coronaviruses cause common colds in humans and important
veterinary diseases 202
A newly emerged coronavirus caused a worldwide epidemic
of severe acute respiratory syndrome (SARS) 202
The SARS coronavirus may have passed from animals to
humans via direct contact 202
Coronaviruses have large, single-stranded, positive sense
RNA genomes 203
Coronaviruses fall into three groups based on genome
sequences 203
Coronaviruses have enveloped virions containing helical
nucleocapsids 204
Coronavirus virions contain multiple envelope proteins 204
Coronavirus spike proteins bind to a variety cellular
receptors 205
The virus envelope fuses with the plasma membrane or an
endosomal membrane 206
The replicase gene is translated from genome RNA into a
polyprotein that is processed by viral proteinases 206
RNA polymerase, RNA helicase, and RNA modifying
enzymes are coded by the replicase gene 207
Replication complexes are associated with cytoplasmic
membranes 207
Genome replication proceeds via a full-length negativestrand
intermediate 208
Transcription produces a nested set of subgenomic
mRNAs 208
Subgenomic mRNAs are most likely transcribed from
subgenomic negative-sense RNA templates 208
The alternative model of discontinuous transcription of
antigenome RNA is unlikely to be correct 209
Assembly of virions takes place at intracellular membrane
structures 211
Adaptability of Coronaviruses 212
20. Paramyxoviruses and Rhabdoviruses 214
The mononegaviruses: a group of related negative-strand
RNA viruses 215
Rabies is a fatal human encephalitis caused by a
rhabdovirus 215
Measles is a serious childhood disease caused by a
paramyxovirus 215
Paramyxovirus and rhabdovirus virions have distinct
morphologies 216
Viral envelope proteins are responsible for receptor binding
and fusion with cellular membranes 217
Genome RNA is contained within helical nucleocapsids 218
Paramyxoviruses enter the cell by fusion with the plasma
membrane at neutral pH 218
Gene order is conserved among different paramyxoviruses
and rhabdoviruses 219
Viral messenger RNAs are synthesized by an RNA
polymerase packaged in the virion 220
Viral RNA polymerase initiates transcription exclusively at
the 3? end of the viral genome 220
The promoter for plus-strand RNA synthesis consists of two
sequence elements separated by one turn of the
ribonucleoprotein helix 220
mRNAs are synthesized sequentially from the 3? to the 5? end
of the genome RNA 222
The P/C/V gene codes for several proteins by using
alternative translational starts and by mRNA
?editing? 223
Functions of P, C and V proteins 224
N protein levels control the switch from transcription to
genome replication 224
Virions are assembled at the plasma membrane 224
21. Filoviruses 226
Marburg and Ebola viruses: sporadically emerging viruses
that cause severe, often fatal disease 227
Filoviruses are related to paramyxoviruses and
rhabdoviruses 228
Filoviruses cause hemorrhagic fever 228
Filovirus genomes contain seven genes in a conserved
order 228
Filovirus transcription, replication and assembly 230
Cloned cDNA copies of viral mRNAs and viral genome
RNA are used for study of filoviruses 230
Multi-plasmid transfection systems allow recovery of
infectious filoviruses 230
Filovirus glycoprotein mediates both receptor-binding and
entry by fusion 231
Ebola virus uses RNA editing to make two glycoproteins
from the same gene 232
Does the secreted glycoprotein play a role in virus
pathogenesis? 232
Minor nucleocapsid proteinVP30 activates viral mRNA
synthesis in Ebola virus 233
Matrix protein VP40 directs budding and formation of
filamentous particles 234
Most filovirus outbreaks have occurred in equatorial
Africa 234
Contents xix
Filovirus infections are transmitted to humans from an
unknown animal origin 235
Spread of filovirus infections among humans is limited to
close contacts 235
Pathogenesis of filovirus infections 236
Clinical features of infection 236
22. Bunyaviruses 238
Most bunyaviruses are transmitted by arthropod vectors,
including mosquitoes and ticks 239
Some bunyaviruses cause severe hemorrhagic fever,
respiratory disease, or encephalitis 240
Bunyaviruses encapsidate a segmented RNA genome in a
simple enveloped particle 240
Bunyavirus protein coding strategies: negative-strand and
ambisense RNAs 240
L RNA codes for viral RNA polymerase 241
M RNA codes for virion envelope glycoproteins 242
S RNA codes for nucleocapsid protein and a nonstructural
protein 243
After attachment via the virion glycoproteins, bunyaviruses
enter the cell by endocytosis 243
Bunyavirus mRNA synthesis is primed by the capped 5? ends
of cellular mRNAs 243
Coupled translation and transcription may prevent
premature termination of mRNAs 244
Genome replication begins once sufficient N protein is
made 244
Virus assembly takes place at Golgi membranes 245
Evolutionary potential of bunyaviruses via genome
reassortment 246
23. Orthomyxoviruses 248
Influenza viruses cause serious acute disease in humans, and
occasional pandemics 249
Influenza virus infections of the respiratory tract can lead to
secondary bacterial infections 249
Orthomyxoviruses are negative-strand RNA viruses with
segmented genomes 249
Eight influenza virus genome segments code for a total of ten
different viral proteins 251
Hemagglutinin protein binds to cell receptors and
mediates fusion of the envelope with the endosomal
membrane 252
M2 is an ion channel that facilitates release of nucleocapsids
from the virion 252
Nucleocapsids enter the nucleus, where mRNA synthesis and
RNA replication occur 253
Capped 5_ ends of cellular pre-messenger RNAs are used as
primers for synthesis of viral mRNAs 254
Viral mRNAs terminate in poly(A) tails generated by
?stuttering? transcription 255
Two influenza A mRNAs undergo alternative splicing in the
nucleus 255
Genome replication begins when newly synthesized NP
protein enters the nucleus 255
Nucleocapsids are exported from the nucleus in a complex
with matrix protein and NS2 256
The NS1 protein interferes with polyadenylation of cellular
mRNAs 256
NS1 also inhibits activation of PKR, an important antiviral
pathway induced by interferon 257
Viral envelope proteins assemble in the plasma membrane
and direct budding of virions 257
Neuraminidase cleaves sialic acid, the cellular receptor that
binds to HA 257
Influenza virus strains vary in both transmissibility and
pathogenicity 257
Genetic variability generates new virus strains that can cause
pandemics 258
The 1918 pandemic influenza A virus was probably not a
reassortant virus 258
Genome sequences from some previous influenza A virus
strains confirm the antigenic shift hypothesis 258
Highly pathogenic influenza A strains in poultry farms could
lead to a new pandemic 259
24. Reoviruses 261
Reoviruses were the first double-stranded RNA viruses
discovered 262
Some members of the Reoviridae are important
pathogens 262
Reoviridae have segmented genomes made of doublestranded
RNA 262
Reovirus virions contain concentric layers of capsid
proteins 263
The attachment protein binds to one or two cellular
receptors 265
During entry, the outer capsid is stripped from virions and
the core is released into the cytoplasm 265
Enzymes in the viral core synthesize and cap messenger
RNAs 266
Translation of reovirus mRNAs is regulated 267
Interferon and PKR: effects on viral and cellular protein
synthesis 267
Synthesis of progeny double-stranded genomes occurs within
subviral particles 268
Reoviruses induce apoptosis via activation of transcription
factor NF- _B 269
Studies of reovirus pathogenesis in mice 270
25. Retroviruses 272
Retroviruses have a unique replication cycle based
on reverse transcription and integration of their
genomes 273
Viral proteins derived from the gag, pol and env genes are
incorporated in virions 273
Retroviruses enter cells by the fusion pathway 274
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Viral RNA is converted into a double-stranded DNA copy by
reverse transcription 275
A copy of proviral DNA is integrated into the cellular
genome at a random site 277
Sequence elements in the long terminal repeats direct
transcription and polyadenylation by host cell
enzymes 277
Differential splicing generates multiple mRNAs 279
The Gag/Pol polyprotein is made by suppression of
termination and use of alternative reading frames 279
Virions mature into infectious particles after budding from
the plasma membrane 280
Acute transforming retroviruses express mutated forms of
cellular growth signalling proteins 281
Retroviruses lacking oncogenes can transform cells by
insertion of proviral DNA near a proto-oncogene 282
26. Human Immunodeficiency Virus
Type 1 284
Human immunodeficiency virus type 1 (HIV-1) and acquired
immunodeficiency syndrome 285
HIV-1 infection leads to a progressive loss of cellular
immunity and increased susceptibility to opportunistic
infections 285
HIV-1 is a complex retrovirus 287
HIV-1 targets cells of the immune system by recognizing
CD4 antigen and chemokine receptors 287
Virus mutants arise rapidly because of errors generated
during reverse transcription 288
Unlike other retroviruses, HIV-1 directs transport of proviral
DNA into the cell nucleus 288
Latent infection complicates the elimination of
HIV-1 289
The Tat protein increases HIV-1 transcription by
stimulating elongation by RNA polymerase II 289
The Rev protein mediates cytoplasmic transport of viral
mRNAs that code for HIV-1 structural proteins 290
Together, the Tat and Rev proteins strongly upregulate viral
protein expression 291
The Vif protein increases virion infectivity by counteracting
a cellular deoxcytidine deaminase 291
The Vpr protein enables the preintegration complex to be
transported to the nucleus 291
The Vpu protein enhances release of progeny virions from
infected cells 291
The Nef protein is an important mediator of
pathogenesis 292
27. Human T-Cell Leukemia Virus
Type 1 294
Discovery of the first human retrovirus 294
Like lentiviruses, HTLV-1 codes for regulatory proteins by
producing doubly-spliced mRNAs 295
HTLV-1 Rex regulates polyadenylation, splicing, and
nuclear export of viral RNAs 296
HTLV-1 Tax regulates transcription of viral and cellular
genes 297
Cell transformation by HTLV-I is mediated by Tax 299
The interleukin-2?autocrine loop stimulates T-cell
proliferation 299
Activation of the Jak-Stat pathway by p12I mimics
interleukin-2 stimulation 299
Cell Cycle Progression: p16INK4A and cyclin-dependent
kinases 300
The mitotic spindle checkpoint and MAD1 300
Downregulation of p53 activity by Tax allows T cell
proliferation 301
Diseases caused by HTLV-1 develop slowly and can be
severe 301
Coinfection by HIV-1 and HTLV-1 is an emerging
problem 302
Antiviral therapy of disease caused by HTLV-1 has not met
with great success 302
28. Hepadnaviruses 303
At least seven distinct viruses cause human hepatitis 304
The discovery of hepatitis B virus 304
Dane particles are infectious virions; abundant noninfectious
particles lack nucleocapsids 305
The viral genome is a circular, partly single-stranded DNA
with overlapping reading frames 305
Nucleocapsids enter the cytoplasm via fusion and are
transported to the nucleus 305
Transcription of viral DNA gives rise to several mRNAs and
a pregenome RNA 307
The roles of hepatitis B virus proteins 308
The pregenome RNA is packaged by interaction with
polymerase and core proteins 309
Genome replication occurs via reverse transcription of
pregenome RNA 309
Virions are formed by budding in the endoplasmic
reticulum 311
Hepatitis B virus can cause chronic or acute hepatitis,
cirrhosis, and liver cancer 312
Hepatits B virus is transmitted by blood transfusions,
contaminated needles, and unprotected sex 313
A recombinant vaccine is available 313
Antiviral drug treatment has real but limited success 313
29 Viroids and Hepatitis Delta Virus 315
Viroids are small, circular RNAs that do not encode
proteins 316
Group A and group B viroids have distinct properties 316
Viroids replicate via linear multimeric RNA intermediates 317
Three enzymatic activities are needed for viroid
replication 317
Contents xxi
How do viroids cause disease? 318
Interaction of viroid RNA with cellular RNAs or proteins
may disrupt cell metabolism 318
Plant satellite RNAs resemble viroids but are
encapsidated 320
Hepatitis delta virus is a human viroid-like satellite
virus 320
Hepatitis delta virus may use two different cellular RNA
polymerases to replicate 320
RNA editing generates two forms of hepatitis delta
antigen 322
Conclusion: viroids may be a link to the ancient RNA
world 322
30. Prions 323
Prions are proteins that cause fatal brain diseases 323
Prion diseases were first detected in domestic
ruminants 324
Human prion diseases can be either inherited or
transmitted 324
The infectious agent of prion diseases contains protein but
no detectable nucleic acid 325
PrPSc is encoded by a host cell gene 325
Differences between PrPC and PrPSc 326
The prion hypothesis: formation of infectious and
pathogenic prions from normal PrPC 326
Is the prion hypothesis correct? 327
Pathology and diagnosis of prion diseases 330
Genetics of prion diseases 330
Prion diseases are not usually transmitted among different
species 330
Strain variation and crossing of the species barrier 331
The nature of the prion infectious agent 331
31. Interferons 333
Virus-infected cells secrete interferons, which protect nearby
cells against virus infection 334
Interferons are a first line of host defense against viruses but
therapeutic use has been limited 334
Interferons _, _, and _are made by different cells and have
distinct functions 335
Transcription of interferon genes is activated by virus
infection or double-stranded RNA 335
Transcriptional activation occurs by binding of transcription
factors to interferon gene enhancers 335
Interferon signal transduction is carried out via the Jak-Stat
pathway 337
Antiviral activities induced by interferon 338
Interferons have diverse effects on the immune system 340
The adaptive immune system 340
Interferons stimulate antigen processing and
presentation 342
Interferon and the development of CD4-positive helper
T-cells 342
The role of interferon in macrophage activation and cellular
immunity 343
Effects of interferons on antibody production 343
Interferons regulate cell growth and apoptosis 343
Viruses have developed numerous strategies to evade the
interferon response 343
Conclusion: interferons are a first line of defense against
virus infection 344
32 Antiviral Chemotherapy 346
The discovery and widespread use of antiviral compounds
began only recently 346
Importance of antiviral drugs for basic science 347
How are antiviral drugs obtained? 347
Targeting drugs to specific steps of virus infection 347
Capsid-binding drugs prevent attachment and entry of
virions 348
Amantadine blocks ion channels and inhibits uncoating of
influenza virions 349
Nucleoside analogues target viral DNA polymerases 349
Acyclovir is selectively phosphorylated by herpesvirus
thymidine kinases 350
Acyclovir is preferentially incorporated by herpesvirus DNA
polymerases 351
Cytomegalovirus encodes a protein kinase that
phosphorylates ganciclovir 351
HIV-1 reverse transcriptase preferentially incorporates
azidothymidine into DNA, leading to chain
termination 354
Nonnucleoside inhibitors selectively target viral replication
enzymes 354
Protease inhibitors can interfere with virus assembly and
maturation 354
Ritonavir: a successful protease inhibitor of HIV-1 that was
developed by rational methods 354
Neuraminidase inhibitors suppress release and spread of
influenza virus 355
Antiviral chemotherapy shows promise for the
future 357
33. Eukaryotic Virus Vectors 358
Many viruses can be engineered to deliver and express
specific genes 358
Virus vectors are used to produce high levels of specific
proteins in cultured cells 359
Gene therapy is an expanding application of virus
vectors 360
Virus vectors are produced by transfection of cells with
plasmids containing deleted genomes 360
Virus vectors are engineered to produce optimal levels of
gene products 361
xxii Contents
ADENOVIRUS VECTORS 362
Adenovirus vectors are widely used in studies of gene transfer
and antitumor therapy 362
Replication-defective adenovirus vectors are propagated in
complementing cell lines 362
Replication-competent adenovirus vectors are useful tools in
antitumor therapy 363
Advantages and limitations of adenovirus vectors 363
RETROVIRUS VECTORS 364
Retrovirus vectors incorporate transgenes into the cell
chromosome 364
Packaging cell lines express retrovirus enzymatic and
structural proteins 364
Strategies for controlling transgene transcription 364
Lentivirus vectors are used for gene delivery to nondividing
cells 365
Production of lentivirus vectors requires additional cis-acting
sequences 366
Applications of retrovirus vectors: treatment of blood
disorders 367
Advantages and limitations of retrovirus vectors 367
ADENO-ASSOCIATED VIRUS VECTORS 367
Adeno-associated virus vectors can insert transgenes into a
specific chromosomal locus 367
Production of AAV vectors usually requires a helper
virus 367
Applications of adeno-associated virus vectors: treatment of
hemophilia 368
Advantages and limitations of AAV vectors 368
34. Viral Vaccines 370
A brief history of viral vaccines 371
Early vaccine technology was crude but effective 372
Embryonated chicken eggs and cell culture played major
roles in recent vaccine development 373
Major categories of viral vaccines 373
Advantages and drawbacks of vaccine types 374
How do viral vaccines work? 375
The role of the immune system in fighting viral
infections 375
Some vaccines target mainly antibody production; others
target the cellular immune response 376
New approaches to vaccine development show great
promise 376
The changing vaccine paradigm 378
Vaccine-associated adverse events 378
Ethical issues in the use of viral vaccines 380
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