Jane Flint is Professor Emerita of Molecular Biology at Princeton University. Dr. Flint's research focused on investigation of the mechanisms by which viral gene products modulate host pathways and antiviral defenses to allow efficient reproduction in normal human cells of adenoviruses, viruses that are used in such therapeutic applications as gene transfer and cancer treatment.

Vincent R. Racaniello is Higgins Professor of Microbiology & Immunology at Columbia University Vagelos College of Physicians & Surgeons. Dr. Racaniello has been studying viruses for over 40 years, including polio- virus, rhinovirus, enteroviruses, hepatitis C virus, and Zika virus. He blogs about virus-es at [...] and is host of This Week in Virology.

Glenn F. Rall is a Professor and the Chief Academic Officer at the Fox Chase Cancer Center, and is an Adjunct Professor in the Microbiology and Immunology departments at the University of Pennsylvania, as well as Thomas Jefferson, Drexel, and Temple Universities. Dr. Rall studies viral infections of the brain and the immune responses to those infections, with the goal of defining how viruses contribute to disease.

Theodora Hatziioannou is a Research Associate Professor at Rockefeller University and is actively involved in teaching programs at Albert Einstein College of Medicine. Dr. Hatziioannou has worked on multiple viruses with a focus on retroviruses and the molecular mechanisms that govern virus tropism and on the improvement of animal models for human disease.

Anna Marie Skalka is a Professor Emerita and former Senior Vice President for Basic Research at the Fox Chase Cancer Center. Dr. Skalka is internationally recognized for her contributions to the understanding of the biochemical mechanisms by which retroviruses replicate and insert their genetic material into the host genome, as well as her research into other molecular aspects of retrovirus biology.

Preface xvii

Acknowledgments xxi

About the Authors xxiii

Key of Repetitive Elements xxv

Part I The Science of Virology 1

1 Foundations 2

Luria's Credo 3

Viruses Defined 3

Why We Study Viruses 3

Viruses Are Everywhere 3

Viruses Infect All Living Things 4

Viruses Can Cause Human Disease 5

Viruses Can Be Beneficial 5

Viruses "R" Us 6

Viruses Can Cross Species Boundaries 6

Viruses Are Unique Tools To Study Biology 6

Virus Prehistory 7

Viral Infections in Antiquity 7

The First Vaccines 8

Microorganisms as Pathogenic Agents 9

Discovery of Viruses 11

The Defining Properties of Viruses 13

The Structural Simplicity of Virus Particles 13

The Intracellular Parasitism of Viruses 13

Cataloging Animal Viruses 18

The Classical System 18

Classification by Genome Type: the Baltimore System 19

A Common Strategy for Viral Propagation 21

Perspectives 21

References 24

Study Questions 24

2 The Infectious Cycle 26

Introduction 27

The Infectious Cycle 27

The Cell 27

Entering Cells 28

Viral RNA Synthesis 29

Viral Protein Synthesis 29

Viral Genome Replication 29

Assembly of Progeny Virus Particles 29

Viral Pathogenesis 29

Overcoming Host Defenses 30

Cultivation of Viruses 30

Cell Culture 30

Embryonated Eggs 35

Laboratory Animals 35

Assay of Viruses 35

Measurement of Infectious Units 35

Efficiency of Plating 38

Measurement of Virus Particles 40

Viral Reproduction: The Burst Concept 49

The One-Step Growth Cycle 49

One-Step Growth Analysis: a Valuable Tool for Studying Animal Viruses 52

Global Analysis 53

DNA Microarrays 54

Mass Spectrometry 56

Protein-Protein Interactions 56

Single-Cell

Virology 56

Perspectives 58

References 59

Study Questions 60

Part II Molecular Biology 61

3 Genomes and Genetics 62

Introduction 63

Genome Principles and the Baltimore System 63

Structure and Complexity of Viral Genomes 63

DNA Genomes 64

RNA Genomes 65

What Do Viral Genomes Look Like? 68

Coding Strategies 69

What Can Viral Sequences Tell Us? 69

The "Big and Small" of Viral Genomes: Does Size Matter? 71

The Origin of Viral Genomes 73

Genetic Analysis of Viruses 74

Classical Genetic Methods 75

Engineering Mutations into Viral Genomes 77

Engineering Viral Genomes: Viral Vectors 83

Perspectives 87

References 87

Study Questions 88

4 Structure 90

Introduction 91

Functions of the Virion 91

Nomenclature 92

Methods for Studying Virus Structure 92

Building a Protective Coat 95

Helical Structures 96

Capsids with Icosahedral Symmetry 99

Other Capsid Architectures 111

Packaging the Nucleic Acid Genome 112

Direct Contact of the Genome with a Protein Shell 112

Packaging by Specialized Viral Proteins 113

Packaging by Cellular Proteins 113

Viruses with Envelopes 115

Viral Envelope Components 115

Simple Enveloped Viruses: Direct Contact of External Proteins with the Capsid or Nucleocapsid 117

Enveloped Viruses with an Additional Protein Layer 118

Large Viruses with Multiple Structure Elements 119

Particles with Helical or Icosahedral Parts 120

Alternative Architectures 123

Other Components of Virions 125

Enzymes 125

Other Viral Proteins 125

Cellular Macromolecules 126

Mechanical Properties of Virus Particles 126

Investigation of Mechanical Properties of Virus Particles 126

Stabilization and Destabilization of Virus Particles 128

Perspectives 128

References 129

Study Questions 130

5 Attachment and Entry 132

Introduction 133

Attachment of Virus Particles to Cells 133

General Principles 133

Identification of Receptors for Virus Particles 135

Virus-Receptor Interactions 137

Entry into Cells 142

Virus-induced Signaling via Cell Receptors 142

Routes of Entry 143

Membrane Fusion 145

Intracellular Trafficking and Uncoating 154

Movement of Viral and Subviral Particles within Cells 154

Uncoating of Enveloped Virus Particles 155

Uncoating of Nonenveloped Viruses 155

Import of Viral Genomes into the Nucleus 159

The Nuclear Pore Complex 159

Nuclear Localization Signals 159

Nuclear Import of RNA Genomes 161

Nuclear Import of DNA Genomes 162

Import of Retroviral Genomes 162

Perspectives 164

References 165

Study Questions 166

6 Synthesis of RNA from RNA Templates 168

Introduction 169

The Nature of the RNA Template 169

Secondary Structures in Viral RNA 169

Naked or Nucleocapsid RNA 170

The RNA Synthesis Machinery 171

Identification of RNA-Dependent RNA Polymerases 171

Three-Dimensional Structures of RNA-Dependent RNA Polymerases 173

Mechanisms of RNA Synthesis 176

Initiation 176

Capping 179

Elongation 179

Functions of Additional Polymerase Domains 181

RNA Polymerase Oligomerization 181

Template Specificity 182

Unwinding the RNA Template 182

Role of Cellular Proteins 183

Paradigms for Viral RNA Synthesis 183

(+) Strand RNA 184

Synthesis of Nested Subgenomic mRNAs 184

(¿) Strand RNA 185

Ambisense RNA 189

Double-Stranded RNA 189

Unique Mechanisms of mRNA and Genome Synthesis of Hepatitis Delta Virus 190

Do Ribosomes and RNA Polymerases Collide? 192

Origins of Diversity in RNA Virus Genomes 193

Misincorporation of Nucleotides 193

Segment Reassortment and RNA Recombination 193

RNA Editing 194

Perspectives 195

References 196

Study Questions 197

7 Synthesis of RNA from DNA Templates 198

Introduction 199

Properties of Cellular RNA Polymerases That Transcribe Viral DNA 199

Some Viral Genomes Must Be Converted to Templates Suitable for Transcription 200

Transcription by RNA Polymerase II 201

Regulation of RNA Polymerase II Transcription 203

Common Properties of Proteins That Regulate Transcription 206

Transcription of Viral DNA Templates by the Cellular Machinery Alone 208

Viral Proteins That Govern Transcription of DNA Templates 209

Patterns of Regulation 209

The Human Immunodeficiency Virus Type 1 Tat Protein Autoregulates Transcription 211

The Transcriptional Cascades of DNA Viruses 217

Entry into One of Two Alternative Transcriptional Programs 226

Transcription of Viral Genes by RNA Polymerase III 230

The VA-RNA I Promoter 231

Inhibition of the Cellular Transcriptional Machinery 232

Unusual Functions of Cellular Transcription Components in Virus-Infected Cells 233

Viral DNA-Dependent RNA Polymerases 233

Perspectives 234

References 235

Study Questions 236

8 Processing 238

Introduction 239

Covalent Modification during Viral Pre-mRNA Processing 240

Capping the 5¿ Ends of Viral mRNA 240

Synthesis of 3¿ Poly(A) Segments of Viral mRNA 243

Internal Methylation of Adenosine Residues 245

Splicing of Viral Pre-mRNA 246

Regulated Processing of Viral Pre-mRNA 249

Editing of Viral mRNAs 255

Export of RNAs from the Nucleus 257

The Cellular Export Machinery 257

Export of Viral mRNA 258

Posttranscriptional Regulation of Viral or Cellular Gene Expression by Viral Proteins 262

Temporal Control of Viral Gene Expression 262

Viral Proteins Can Inhibit Cellular mRNA Production 264

Regulation of Turnover of Viral and Cellular mRNAs in the Cytoplasm 266

Intrinsic Turnover 266

Regulation of mRNA Stability by Viral Proteins 267

mRNA Stabilization Can Facilitate Transformation 267

Nonsense-Mediated mRNA Decay 267

Noncoding RNAs 271

Small Interfering RNAs and Micro-RNAs 271

Long Noncoding RNAs 276

Circular RNAs 278

Perspectives 278

References 279

Study Questions 281

9 Replication of DNA Genomes 282

Introduction 283

DNA Synthesis by the Cellular Replication Machinery 284

Eukaryotic Replicons 284

Cellular Replication Proteins 287

Mechanisms of Viral DNA Synthesis 287

Lessons from Simian Virus 40 288

Replication of Other Viral DNA Genomes 290

Properties of Viral Replication Origins 294

Recognition of Viral Replication Origins 296

Viral DNA Synthesis Machines 301

Resolution and Processing of Viral Replication Products 301

Exponential Accumulation of Viral Genomes 302

Viral Proteins Can Induce Synthesis of Cellular Replication Proteins 303

Synthesis of Viral Replication Machines and Accessory Enzymes 304

Viral DNA Replication Independent of Cellular Proteins 304

Delayed Synthesis of Structural Proteins Prevents Premature Packaging of DNA Templates 305

Inhibition of Cellular DNA Synthesis 305

Synthesis of Viral DNA in Specialized Intracellular Compartments 305

Limited Replication of Viral DNA Genomes 308

Integrated Parvoviral DNA Can Be Replicated as Part of the Cellular Genome 308

Different Viral Origins Regulate Replication of Epstein-Barr Virus 310

Limited and Amplifying Replication from a Single Origin: the...