The Molecular Biology of Poliovirus

I: The Poliovirus.- 1. History.- I. Introduction.- II. Early History-Occasional Nonepidemic Cases of Poliomyelitis.- III. The 18th and 19th Centuries: Polioepidemics, Poliomyelitis Is Described as a Clinical Entity.- IV. Early 20th Century: Research on Polio Begins, Virology Is still a Clinical Discipline.- V. The Development of Vaccines Against the Poliomyelitis Virus.- VI. The Fifties and the Advent of Molecular Biology.- VII. Advances in Polio Research During the Past Two Decades.- 2. Classification.- I. General Considerations.- A. Nature of the Genome.- B. The Cryptogram.- II. The Distinguishing Features of Picornaviruses.- III. Comparison of Different Picornavirus Genera.- A. Disease Aspects.- B. Serotypes.- C. Physical Properties.- D. Relative Relatedness to Polioviruses.- 3. Composition and Structure of the Virion.- I. Introduction.- II. Composition and Physical Properties of the Virion.- III. Overall Architecture of the Viral Capsid.- A. Building Blocks.- 1. Functional Requirements.- 2. Biological Arguments of Building Block Economy and Efficiency of Assembly.- 3. Thermodynamic Forces and Structural Consequences.- B. The Geometric Design: The Icosahedron.- 1. Helical Tubes Versus Spherical Shells.- 2. Cubic Symmetries, the Platonic Polyhedra.- 3. The Icosahedron.- a) Advantages of Icosahedral Symmetry.- b) Limitations of the Icosahedral Skeleton as a Model for Virus Structure.- c) Related Regular Polyhedra with Icosahedral Symmetry.- d) The Bonding Pattern of an Icosahedral Lattice.- C. Experimental Evidence that the Poliovirus Capsid Is an Icosahedral Lattice.- 1. X-Ray Diffraction Studies.- 2. Electron Microscopic Observations.- 3. Resolving the 32 or 60 Capsomers Controversy.- 4. Similar Construction Principles for Picornaviruses and the Small Plant Viruses.- IV. Characterization of the Building Blocks: The Capsid Proteins.- A. Separation and Identification.- B. Amino Acid Composition and Sequence.- C. Microheterogeneity.- V. Relative Localization of the Viral Proteins in the Capsid and Bonds Involved.- A. General Reflections.- 1. Geometric Restrictions.- 2. Structural Principles Borrowed from Plant Picornaviruses.- 3. Implications for the Capsid Features of Poliovirus: The Concept of a Rigid Capsid Backbone and Variable Surface Protrusions.- B. Experimental Results.- 1. Specific Chemical Modifications of Capsid Proteins.- 2. Chemical Crosslinking of Capsid Proteins.- 3. UV Irradiation of Poliovirus.- VI. The Dissociation of Poliovirus.- A. Breakdown During Preparatory Procedures for Electron Microscopy.- B. Alkaline Degradation.- C. Heat Degradation.- D. Guanidine Degradation.- E. Urea Degradation.- F. Reassociation of Poliovirus from Products of Urea Degradation.- G. Conclusions.- VII. Conformational Forms of the Poliovirus Capsid.- A. Poliovirus Capsid Structures During Early Interactions with the Host Cell and upon Experimentally Induced Disruption: Two Conformational States of the Shell.- B. Viral Structures During Maturation.- C. Two Distinct, Reversibly Interconvertible Conformational States of Intact Poliovirions.- D. Conformational Alterations Monitored by Changes in the Intrinsic and Induced Fluorescence of Poliovirus Components.- E. Dense Particles.- F. Possible Functions of the Alternate Conformational States of the Poliovirus Capsid.- VIII. Antigenicity.- A. The Main Poliovirus Antigenic Sites: The D- and C-Antigenic States.- 1. Attempts to Identify the C- and D-Antigenic Determinants in Terms of the Constituent Capsid Proteins with Polyclonal Antisera.- 2. Studies with Monoclonal Antibodies.- B. Other, Minor Antigenic Sites on Poliovirus.- C. Serodifferentiation of Polioviruses.- IX. Summary.- A. Composition.- B. Structure.- C. The Capsid Proteins.- 1. Number of Peptides and Type of Association on the Structure Units.- 2. Relative Localizations.- 3. Heterogeneity.- D. Two Conformational States of the Capsid.- E. Antigenicity.- 4. Structure and Function of the Genome.- II. Genome Structure.- A. Characteristic Features.- B. Structural Organization.- C. Secondary Structure.- D. The Genome-Linked Protein VPg.- III. Nucleotide Sequences.- A. Oligonucleotide Mapping.- B. Cloning of Poliovirus cDNA.- C. The Consensus Sequence of Poliovirus Type I.- D. Terminal Sequences.- 1. 5? End Sequences.- 2. 3? End Sequences.- 3. The Poly (A) Tract.- E. Features of the Coding Region.- 1. Codon Usage.- 2. Cleavage Signals for Proteolytic Processing.- IV. Genome Products and Gene Order.- A. Mapping of the Genome Products on the Polioviral Genome.- 1. Relatedness of Viral Proteins: Tryptic Peptide Analysis.- 2. Biochemical Mapping.- 3. The Genome Map as Deduced from Genetic Studies.- B. Function of the Viral Proteins.- 1. The P-1 Proteins: Coat-Proteins, Proteinkinase, Shut-Off of Protein Synthesis.- 2. The P-2 Proteins: Guanidine Sensitivity, VP0 Protease.- 3. The P-3 Proteins: glu/gly Protease, VPg Replicase, Cytophathic Effect.- V. Genomic Variation of Polioviruses.- VI. Genetics.- A. Mutations.- B. Mutant Types.- 1. Temperature Sensitive Mutants.- 2. Structural Markers.- 3. Non Structural Markers.- C. Genetic Recombination.- D. Mechanism of Recombination.- VII. Summary.- II: The Replication of Poliovirus.- 5. Introduction.- I. The "Life-Cycle" of Poliovirus.- II. Timecourse of Poliovirus Replication.- III. The Host Cell.- A. Constituents of the Cell.- B. The Nucleus.- C. The Plasma Membrane.- D. The Ionic Environment.- E. The Cytoskeleton.- F. Intracellular Membranes.- G. The Cell Cycle.- H. Employment of the Metabolic Machinery of the Host Cell by the Virus.- IV. Some Speculations on Abortive Infections of Poliovirus..- 6. Morphological Alterations of the Host Cell as an Essential Basis for Poliovirus Replication.- I. Introduction.- II. Microscopic Oberservations.- A. Light Microscopy.- B. Electron Microscopy.- 1. The Nucleus.- 2. Ribosomes.- 3. "Viroplasm".- 4. Alterations of Intracellular Membranes.- a) Nuclear "Extrusions".- b) Membraneous Cisternae.- c) Biochemical Aspects of Membrane Formation.- 5. Changes in the Cytoskeletal Framework.- 6. Assembly and Release of Progeny Virions.- 7. Lysosomes and Autophagic Vesicles.- 8. Inhibitors of Morphological Alterations.- III. Speculations on the Function of Compartmentalization in Virion RNA Synthesis and Assembly.- A. Concerning the Mode of Vesicle Formation.- B. Concerning the Relative Localization of RNA Synthesis and Virion Assembly with Respect to the Vesicle Membrane.- IV. Poliovirus induced Alterations in Functions of the Plasma Membrane, in the Intracellular Ionic Environment, and in Cell Size.- A. Membrane Changes Accompanying Adsorption and Penetration of Poliovirus.- B. Membrane Alterations Accompanying Virus Replication at the Maximal Rate.- V. Summary.- 7. Early Interactions of Virus and Host Cell.- I. Introduction.- A. Overview and Definition of Terms.- B. The Superposition of Abortive and Productive Pathways in Infection.- II. Adsorption and Attachment.- A. The Reaction Partners.- 1. The Virus Particle.- 2. The Virus Receptor Complex on the Host Cell.- a) Properties of the Poliovirus Receptor.- b) Other Functions and Components of the Receptor Complex.- c) Number of Attachment Sites/Cell.- d) Specificity and Genetics of Virus Receptors.- B. The Interaction of Poliovirions with the Host Cell Membrane.- 1. Adsorption.- 2. Attachment.- 3. Response of the Plasma Membrane.- a) Changes in Membrane Fluidity and Capping of Viruses.- b) Changes in Membrane Permeability and Membrane Potential.- c) Interaction with Modifying and Stabilizing Membrane Components.- III. Penetration of Virus Particles into the Cell: Insertion and Phagocytosis.- IV. Uncoating: A Multistep Process.- A. Possible Steps and Sites of Uncoating.- B. The Fate of the Parental Capsid Proteins.- V. Infection of Cells Lacking Receptors.- A. Introduction.- B. Adsorption of Viral RNA to Cells.- C. Penetration of Isolated Viral RNA into Cells.- 1. RNA-Penetration by Passive Influx of RNA.- 2. Stimulation of Active Uptake of Viral RNA.- 3. Entrance of Poliovirus RNA into Cells Via Lipid Vesicles (Liposomes).- D. Cellular Competence for Infection by Viral RNA.- 1. Optimal Conditions for the Use of Polycations.- 2. Relationship Between RNA Concentration and Yield of Infections Centers.- 3. The Combined Effect of Dimethylsulfoxide and DEAEDextran on the Competence of Cells for Infection by Viral RNA.- 4. Competence of HeLa Cells for Infection by Viral RNA at Different Stages in a Cell Growth Cycle.- E. Conclusions.- VI. Summary.- 8. Translation of the Viral Genome.- I. Mammalian Protein Synthesis.- A. The Protein Synthesizing Machinery.- 1. Ribosomes.- a) Structure and Composition.- b) Monosomes and Polysomes.- c) Free and Membrane Bound Polysomes.- 2. mRNA.- a) The Cap.- b) The 5? Terminal Untranslated Region.- c) The 3? Terminal Untranslated Region and the Poly A Tract.- d) Monocistronic mRNAs and Potential Internal Initiation Sites.- e) mRNPs.- 3. Initiation Factors and the Process of Initiation.- 4. Elongation and Termination of Translation.- 5. Cotranslational Processing and Membrane Insertion of Nascent Polypeptide Chains.- B. The Regulation of Protein Synthesis.- 1. The Role of Culture Conditions.- 2. Competition Between mRNAs.- a) The Role of mRNA Concentration.- b) Relative Translational Efficiencies of mRNAs.- c) The Role of Limiting Initiation Components.- 3. Alteration or Inactivation of the Cap Binding Protein and Other Initiation Factors.- 4. Modification of Ribosomes.- 5. The Role of Uncharged tRNA.- 6. Transfer of mRNAs Between Untranslatable and Translatable Pools.- 7. Control of Free and Membrane Bound Pools of Ribosomes.- II. Translation of the Poliovirus Genome.- A. Overview and Introduction.- B. Translation of Poliovirus RNA in Cell Free Extracts.- 1. General Comments.- 2. Initiation of in vitro Translation.- 3. In vitro Elongation and Termination.- C. The Additional Complexity of in vivo Translation During Infection.- 1. The Shut-Off Phenomenon.- a) The Activation of an Inherent Host-Cell Regulatory Mechanism.- b) Competition Between Viral and Host Cell mRNAs.- c) The Role of Ionic Disturbances and Membrane Leakiness.- d) Alterations of Initiation Factors or Ribosomes.- e) The Role of Virus-Specific Factors in Mediating the Shut-Off.- f) In Summary, a Concert of Mechanisms with a Purpose.- 2. Non-Uniform Synthesis of Viral Proteins.- 3. The Role of Interaction Between Poliovirus Proteins and Intracellular Membranes.- 4. Distribution of Viral Proteins.- 5. Protein Processing.- a) The Role of Cleavage.- b) Types of Cleavages.- c) Types of Proteases.- d) Role of Cleavage in RNA-Replication.- e) Interference with Protein Processing.- f) The Effect of Guanidine on the Processing of Polioviral Proteins.- III. Summary.- 9. Replication of the Viral RNA.- I. Introduction.- II. Isolation and Characterization of Virus Specific RNAs Isolated from Infected Cells.- A. Preparation and Purification of Poliovirus Specific RNAs.- B. Properties of Poliovirus Specific RNAs.- 1. The Single-Stranded Viral RNA.- 2. The Replicative Form-RNA.- 3. The Replicative Intermediate-RNA.- 4. Double Stranded Forms of RNA-Extraction Artefacts?.- III. Time Course and Kinetics of Synthesis of Virus Specific RNAs.- A. The Onset of RNA Synthesis.- B. The Exponential Phase: cRNA ? mRNA ? cRNA.- C. The Linear Phase: cRNA ? vRNA, mRNA.- D. Cessation of RNA Synthesis.- IV. The Sites of RNA Synthesis.- V. The Viral RNA-Polymerases.- A. The Crude Replication Complex: Synthesis of Plus Strand RNA.- B. Soluble Replicase(s): Synthesis of Minus Strand RNA.- VI. The Effects of Guanidine on Poliovirus Replication.- VII. Some Thoughts on the Mode of RNA Replication.- A. Initiation of Viral RNA Synthesis.- B. Elongation of Viral RNA Replication.- C. Inhibition of Host Nuclear Functions.- D. On the Infectivity of RF-RNA.- E. Regulation of Viral RNA Synthesis.- VIII. Summary.- 10. Assembly of the Virion.- I. The Cytoplasmic Sites of Assembly-Virus-Induced Intracellular Membranes.- A. Electron Microscopic Observations.- B. Biochemical Approaches.- II. Subviral Particles in the Infected Cell-Potential Assembly Intermediates.- A. Overview.- B. NCVP1a and the 5S Protomer.- C. The 14S Pentamer.- D. The 55S Particle.- E. The 80S Shell.- F. Ribonucleoprotein Particles.- 1. The Slow Sedimenting (80S) RNPs.- 2. The 125S and 150S Provirion(s).- 3. Association of RNPs with the Replication Complex in Smooth Membranes.- III. Assembly Kinetics.- A. Chasing of Radioactive Precursors: Nucleosides and Amino Acids.- B. Assembly of Isolated Subviral Particles in vitro.- 1. Self Assembly of Isolated Subunits.- 2. Assembly-Enhancing Activity in Extracts of Infected Cells.- C. Studies with Inhibitors of Assembly.- 1. Reversible Inhibition of Assembly by Py-11.- 2. Studies with the Assembly Inhibitor Guanidine.- 3. Inhibition of Poliovirus Maturation Under Hypotonic Culture Conditions.- 4. Assembly Defective Mutants.- IV. The Individual Steps of Assembly.- A. Principles of Assembly.- B. Formation, Activation, and Assembly of the 5S Protomer.- C. Activation and Assembly of the 14S Pentamer.- D. Encapsidation of the Viral RNA.- 1. Condensation of the Viral RNA.- 2. Formation of the RNP.- a) Assembly Around an RNP Core.- b) Insertion of RNA into a Procapsid.- 3. Stepwise Condensation of the RNPs.- 4. The Possible Role of Mg++.- E. The Final Morphogenetic Cleavage.- V. Summary.- 11. Conclusions.- Appendix I: Laboratories Engaged in Poliovirus Research.- Appendix II: Poliovirus Models.- A. A Paper Model of a Prototype Picornavirus.- B. The Apple Model of Poliovirus.- Appendix III: The Geometry of Isometric Polyhedra.- A. The Platonic Polyhedra.- a) Models.- b) Characteristics.- c) Duality.- d) The Golden Proportion.- e) Geometric Restriction of the Maximal Number of Subunits.- B. Other Icosahedron-Related Polyhedra.- a) Characteristics.- b) The Triangulation-Number-Classification of Icosahedral Lattices.- Appendix IV: Complete Nucleotide and Amino Acid Sequences of Poliovirus Type 1, 2 and 3.- References.