Fundamentals of Virology

Posted on May 28, 2025 in Biology

Introduction to Virology

  • Comparing Viruses and Cellular Life

    • Viruses are infectious, obligate intracellular parasites, while cellular life can reproduce independently.

    • Viruses possess either a DNA or RNA genome, while cellular life possesses a DNA genome.

    • Viruses require a host cell for ribosomes, energy sources, genome replication, assembly, and preformed components.

  • Ubiquity of Viruses

    • Viruses are everywhere and infect all life forms.

    • There are more viruses in a liter of seawater than humans on the planet.

  • Importance of Virology for Health and Economics

    • Viruses cause diseases, such as those caused by caliciviruses and herpesviruses.

    • The study of viruses has led to the development of mRNA vaccines.

  • Virology’s Insights into Host Biology

    • Viruses use and abuse cell machinery, providing insights into fundamental principles in biology, such as replication, transcription, processing, translation, and immune responses.

  • Exploiting Viruses as Medical Tools

    • Viruses are used in gene therapy.

    • Viruses are used to deliver genes into T cells to better recognize and kill cancer cells.

The Viral Infectious Cycle

  • Methods for Growing Viruses

    • Viruses can be grown in living hosts (plants or animal models), eggs, or cultured cells.

    • Cultured cells can be primary cells (limited generations, diploid) or continuous cell lines (unlimited generations, aneuploid).

  • Measuring Virus Levels

    • Biological assays (measure infection efficiency): Plaque assay, endpoint dilution, and transformation.

    • Physical measurements (require no infection): Hemagglutination, counting particles by electron microscopy, and flow virometry.

  • Understanding Virus Growth Measurement

    • Virus growth measurement is a useful tool to study kinetics of replication and compare fitness of viruses.

  • Calculating Viral Titer

    • Titer = # plaques / (volume in ml * dilution factor).

    • Units are Plaque-Forming Units (PFU) per ml.

  • Multiplicity of Infection (MOI)

    • MOI is the ratio of viruses added to host cells present.

    • A high MOI means most or all cells are infected.

  • Diagnosing Viral Infections

    • By testing for the viral genome (RNA or DNA), viral proteins (antigen), or antibodies to the virus.

    • Methods include reverse transcription and PCR.

Viral Genomes and Genetics

  • Baltimore Classification Scheme

    • The Baltimore classification system categorizes viruses based on their genome type and how mRNA is produced.

  • mRNA Transcription from Viral Genomes

    • dsDNA viruses are ready to be transcribed.

    • (+) RNA viruses can be directly translated by host ribosomes.

    • (-) RNA viruses need to be converted to (+) RNA by viral polymerases before being translated.

  • Enzymes for Viral Genome Replication

    • DNA viruses use DNA-dependent DNA polymerase (DdDP) for genome replication.

    • RNA viruses use RNA-dependent RNA polymerase (RdRP) for genome replication.

  • Generating and Using Viral Mutants

    • Mutants can be experimentally induced (UV, chemical mutagens, directed genetic engineering) or spontaneously generated.

    • Mutants are used to map phenotype to genotype and study extinct viruses.

Viral Structure

  • Basic Components of a Virion

    • The basic components are the nucleic acid genome, capsid, and envelope.

  • Functions of Virion Components

    • The genome carries the genetic information. The capsid protects the genome. The envelope helps in attachment and entry.

  • Shapes of Virus Particles

    • The shapes are helical (rod-shaped) and icosahedral (spherical).

  • Determining Virus Structure

    • Methods include electron microscopy (transmission EM, cryo-EM), atomic force microscopy, X-ray crystallography, and biochemical characterization.

Viral Attachment and Host Cell Entry

  • Host Cell Receptor Types for Viruses

    • Viruses use sugars on proteins, sugars on lipids, and proteins as receptors.

  • Viral Entry Pathways by Structure

    • Enveloped viruses enter by fusion at the plasma membrane or through endocytosis.

    • Non-enveloped viruses enter via endocytosis.

  • Steps in Viral Entry

    • Entry involves attachment, internalization, and membrane rupture/fusion.

  • Host and Environmental Factors in Entry

    • pH and cellular proteases disrupt membranes.

  • Nuclear Entry Mechanisms for Viruses

    • Some viruses reach the nucleus via nuclear localization signals, ejection, or uncoating and release.

RNA Synthesis from RNA Templates

  • mRNA and Genomic RNA Synthesis in RNA Viruses

    • (+) RNA viruses use their genome as mRNA.

    • (-) RNA viruses use RdRp to synthesize mRNA and antigenome.

  • Mechanisms of Viral RNA Synthesis Regulation

    • Initiation can be primer-independent (de novo) or primer-dependent (protein or RNA primer).

    • Regulation involves different RdRp, nucleoproteins, and stop-start transcription.

  • RNA Virus Replication and Genetic Diversity

    • RNA replication increases viral diversity through error-prone replication, recombination, and reassortment.

Viral Transcription Processes

  • Requirements for DNA Virus RNA Synthesis

    • Requirements include template, primer, and enzyme.

  • Advantages of Temporally Regulated Viral Transcription

    • Temporal regulation coordinates viral building block production.

  • Regulatory Effects of Viral Transcription Components

    • Viral transcription factors can cooperate with cellular proteins to transcribe certain viral genes and repress others.

  • Regulation of Viral Latency

    • Viral latency is regulated by latency-associated transcripts (LATs).

Viral RNA Processing

  • Eukaryotic Virus mRNA Processing

    • Eukaryotic virus mRNA is processed by splicing, capping, and polyadenylation.

  • Location of mRNA Synthesis and Processing

    • mRNA synthesis and processing occur in the nucleus for DNA viruses and in the cytoplasm for RNA viruses.

  • Modulating Viral mRNA Half-Life

    • Viral strategies can tip the balance to favor the virus and keep host mRNAs out of the cytoplasm.

Viral DNA Replication

  • DNA Virus Replication Location and Protein Sources

    • Large DNA genomes may encode their own DNA replication factors.

  • Requirements for Viral DNA Synthesis

    • DNA synthesis requires DdDP, template-dependent synthesis, and a primer.

  • DNA Replication Modes: Fork vs. Strand Displacement

    • Some dsDNA viruses: RNA primer always required, primer synthesized by host (SV40) or viral primase (HSV-1).

    • ssDNA and dsDNA viruses: RNA primer never required, DNA or protein primer.

  • Solving the ‘End Problem’ in DNA Replication

    • Strategies include circularization, protein primer, and self-priming.

Reverse Transcription and Viral Integration

  • Reverse Transcriptase and the Central Dogma

    • Reverse transcriptase (RT) challenges the central dogma by converting RNA to DNA.

  • Process and Requirements of Reverse Transcription and Integration

    • Reverse transcription requires RT, a template, and a primer.

    • Integration is mediated by viral integrase.

  • Reverse Transcriptase and Viral Diversity

    • RT has no editing/proofreading activity, leading to high mutation rates and viral diversity.

  • Examples of Reverse Transcriptase Users

    • Examples include retroviruses and hepadnaviruses.

Viral Translation Mechanisms

  • Eukaryotic Translation Initiation: Dependent and Independent

    • 5’ end-dependent initiation involves eukaryotic initiation factors (eIFs) and GTP.

    • 5’ end-independent initiation involves internal ribosome entry sites (IRES).

  • Viral Strategies for Expanding Coding Capacity

    • Strategies include internal initiation for second ORFs, translation reinitiation by ribosome shunting, and template circularization.

  • Inhibition of mRNA Translation by Viruses and Host Cells

    • Viruses restrict cellular gene expression by targeting mRNA.

    • Host cells inhibit translation via stress granules and P-bodies.

  • Viruses Counteracting Host Translation Defenses

    • Viruses counteract PKR by producing decoys for the kinase and using short dsRNA.

Viral Assembly and Host Cell Escape

  • Virus Exit Strategies by Virion Structure

    • Non-enveloped viruses are usually released via cell lysis.

    • Enveloped viruses exit by budding through host membranes.

  • Viral Protein Localization and Assembly

    • Viral proteins are trafficked to the assembly site via nuclear localization signals.

  • Principles of Capsid and Nucleocapsid Self-Assembly

    • Capsid/nucleocapsid assembly can be concerted (genome packaged as capsid assembles) or sequential (genome packaged into preformed capsid).

  • Mechanisms Driving Viral Release

    • Viral release is driven by budding through host membranes.