Lecture Three: Virion Structure and Viral Genetics

• A virion is a complete virus particle with one or more DNA or RNA molecules enclosed in a protein coat.
• Many viruses have segmented genomes, for example, influenza and rotavirus.
• Some viruses have additional layers, such as a lipid membrane.
• Viruses cannot reproduce independently of host cells and must direct mRNA synthesis to produce proteins.
• Viral genomes do not encode a complete translation system, so viral protein synthesis depends on the host cell’s translational machinery.
• The origin of viruses is unclear because they do not fossilize, making it difficult to study their deep evolutionary history.
• Insights into viral history are inferred from genetic sequence data.
• Viral genomes exhibit mosaicism, meaning they are a mix of genes from different origins.
• Genomic studies suggest that viruses have no single origin.
• Viruses show "modularity," meaning genes and gene families have independent origins.
• Genetic studies use colors to indicate genetic similarities across different viruses.

Hypotheses for the Origin of Viruses

• The regressive hypothesis proposes that viruses are degenerate forms of intracellular parasites.
• These viruses evolved from more complex, free-living organisms that lost genetic material.
• Genome reduction is similar to that seen in Nanoarchaeum equitans, a symbiotic archaeon.
• The progressive hypothesis suggests viruses originated as fragments of genetic material that escaped from cells.
• DNA viruses evolved from plasmids or transposable elements, later gaining coat proteins and transmissibility.
• Retroviruses evolved from retrotransposons, while RNA viruses originated from mRNA.
• The coevolution hypothesis proposes that viruses co-evolved with early life forms.
• Viral evolution may date back to the RNA world.
• The multiple origins theory suggests all three hypotheses may be correct and explain different types of viruses.
• Evidence suggests viruses have multiple origins.
• Free-living bacteria may have devolved into viruses, supporting the regressive hypothesis.
• Evolution does not always progress in one direction, as seen in cave fish losing eyesight.

Progressive Hypothesis: Viruses and Retrotransposons

• Retroviruses (e.g., HIV) and retrotransposons behave similarly, except retroviruses have an extracellular stage.
• Retrotransposons make up approximately 42% of the human genome.

Key Features of Retrotransposons

• The Gag gene produces proteins that form virus-like particles in the cytoplasm where reverse transcription occurs.
• The Pol gene produces three proteins: protease, reverse transcriptase, and integrase.
• The Env (envelope) gene produces a glycoprotein forming the capsid and envelope of viruses.

Viral Envelopes

• Many viruses have an outer flexible membranous layer called the envelope.
• Animal virus envelopes (lipids and carbohydrates) usually arise from the host cell plasma or nuclear membranes.
• Envelope proteins facilitate attachment to host cell receptors.

Virus Life Cycle

• The five steps of viral infection are attachment, entry, synthesis, assembly, and release.
• Attachment involves the virus binding to a host cell receptor.
• Entry involves the virus entering the host cell via membrane fusion or endocytosis in eukaryotic cells.
• Entry in bacteriophages happens via tail penetration.
• Synthesis involves the viral genome directing host machinery to produce viral components.
• Assembly involves new virions being assembled inside the host cell.
• Release involves virions exiting the host cell through lysis or budding.
• A one-step growth curve demonstrates progression through the virus life cycle.

Virus Entry Mechanisms of Eukaryotic Viruses

• Membrane fusion is one method of entry.
• Receptor-mediated endocytosis is another entry mechanism.

Main Ways Viruses Enter Host Cells

• Fusion with the Plasma Membrane (Enveloped Viruses): The viral envelope directly fuses with the host cell membrane, releasing the viral capsid and genome into the cytoplasm, for example, HIV and influenza.
• Endocytosis (Enveloped Viruses): The virus binds to a receptor, triggering the host cell to engulf it in a vesicle. Inside the cell, the vesicle acidifies, causing the viral envelope to fuse with the vesicle membrane and release the genome, for example, hepatitis C and dengue virus.
• Endocytosis (Non-Enveloped Viruses): The virus is taken up into a vesicle by receptor-mediated endocytosis. Once inside, the acidic environment or viral proteins break open the vesicle, releasing the genome into the cytoplasm, for example, adenovirus and poliovirus.
• Bacteriophages use tails to penetrate host membranes and inject their genomes.
• Virus self-assembly is a spontaneous process, with viral proteins naturally assembling into complete viruses once synthesized.
• Protein and nucleic acid affinities, along with shape and surface charge interactions, guide proper assembly.
• Some viruses form capsids first, packaging the genome inside, using the procapsid model.
• Others assemble capsids around the genome as they form.
• Cells do not use double stranded RNA, so viruses protect it from host destruction.

Virion Release

• A nucleocapsid is a viral genome enclosed within a protein capsid before it gains an envelope.
• A virion is a fully assembled, infectious virus particle, which may include an envelope.
• Hemagglutinin (HA) is a viral surface protein that facilitates attachment to host cells, particularly in influenza viruses.
• Influenza virus release involves viral envelope proteins inserted into the host cell membrane.
• Matrix proteins line the inner surface of the host membrane.
• Nucleocapsids are transported to the membrane via host microtubules.
• The plasma membrane protrudes, enclosing nucleocapsids.
• The protruding membrane pinches off, forming a mature virion that can infect new cells.

Bacteriophage Lysis: Breaking Free

• Gram-positive bacteria have thick peptidoglycan and no outer membrane, requiring degradation of peptidoglycan for lysis.
• Gram-negative bacteria have thin peptidoglycan and an outer membrane, requiring breaching of both layers for lysis.
• Holin creates holes in the bacterial inner membrane, controls timing of lysis, and allows endolysins to reach the peptidoglycan.
• Endolysin is an enzyme that cuts peptidoglycan, destroying the bacterial wall.
• It works alone in Gram-positives but needs extra help (e.g., spanins) in Gram-negatives to disrupt the outer membrane.
• Holins act like a self-destruct countdown for bacteria.
• Endolysins tear apart bacterial walls with precision.
• This represents a perfectly timed escape strategy evolved by bacteriophages.

Viruses in the Human Genome

• The human genome has a total size of about 3 billion base pairs (bp).
• Protein-coding genes comprise about 20,000 (~10 million bp).
• Most of the genome does not code for proteins but may have regulatory or structural roles.
• "Old" genes can be repurposed for new functions.
• Viruses have contributed to evolutionary innovations.
• Syncytin is a viral protein essential for placental development, expressed in syncytiotrophoblast, the outer layer of the placenta.
• It enables the fusion of cytotrophoblast cells into a syncytial layer.
• Without syncytin, mammalian pregnancy would not be possible.
• Syncytin is homologous to Env proteins in retroviruses (e.g., HIV).
• Retroviral Env proteins allow virus fusion with host cells.
• In humans, syncytin is encoded by ERVWE1, an endogenous retroviral element.
• Gag and Pol genes are inactivated, but Env is retained for placental function.
• Proto-mammals first captured syncytin from a virus to form placentas.
• Different mammalian lineages swapped out earlier syncytin genes for improved versions over time.
• At least eight independent viral captures contributed to placental evolution.
• Cancer is a complex, multistep process involving genetic and environmental factors.
• Oncogenes are genes that, when mutated or overexpressed, drive uncontrolled cell growth.
• Viruses contribute to carcinogenesis, with approximately 20% of cancers linked to viruses.
• Viral genes disrupt cell cycle regulation.
• Some viruses insert oncogenes into the host genome.
• Chronic infections cause inflammation, increasing mutation rates.
• Human Papillomavirus (HPV) is linked to cervical, anal, and throat cancers.
• Some viruses establish persistent infections, remaining in the host for years or a lifetime.
• Latent viruses are dormant within cells, reactivating under stress (e.g., Herpes, HIV).
• Chronic viruses continuously replicate at low levels (e.g., Hepatitis B & C).
• Long-term viral infections can lead to cancer by integrating into host DNA, disrupting cell cycle control (e.g., HPV, Epstein-Barr, Hepatitis B & C).

Mechanism of HPV Oncogenesis

• pRb (Retinoblastoma Protein) binds to E2F transcription factors to prevent premature cell cycle progression.
• During the late G1 phase, cyclin-dependent kinases (CDKs) phosphorylate pRb, releasing E2F and allowing transcription of genes for S phase.
• HPV E7 mimics CDKs, binding to pRb and preventing it from inhibiting E2F.
• Free E2F drives uncontrolled transcription of genes needed for S phase progression.
• This leads to unscheduled cell cycle progression and uncontrolled proliferation, increasing the risk of cancer.
• Normal pRb function ensures controlled cell cycle entry into S phase.
• HPV E7 bypasses this regulation, causing continuous cell division and increasing the likelihood of mutations.
• This cellular dysregulation contributes to oncogenesis, making HPV a major cause of cervical and other cancers.

Human Cell Cycle

• G1 Phase (Gap 1): Cell grows, produces proteins, and prepares for DNA replication; a checkpoint ensures the cell is ready for DNA synthesis.
• S Phase (Synthesis): DNA replication occurs; each chromosome duplicates into sister chromatids.
• G2 Phase (Gap 2): Cell prepares for mitosis, producing necessary proteins; a checkpoint ensures DNA replication is complete and undamaged.
• M Phase (Mitosis): Division of the nucleus into two identical daughter cells through prophase, metaphase, anaphase, telophase, and cytokinesis.
• G0 Phase (Resting State): Cells that do not divide (e.g., neurons) enter this phase indefinitely.

Overview of Viral Replication Mechanisms

Group I: dsDNA Viruses

• Viral DNA is transcribed into mRNA by DNA-dependent RNA polymerase (DdRP), either host or virus-encoded.
• mRNA is translated into viral proteins by host ribosomes.
• dsDNA replicates using DNA-dependent DNA polymerase (DdDP), either from the host or virus.
• Newly synthesized dsDNA is packaged into viral capsids for progeny virus formation.

Group II: ssDNA Viruses

• Host DNA polymerase converts ssDNA into dsDNA.
• Cells need a primer; ssDNA self-primes by forming a hairpin loop.
• dsDNA serves as a template for mRNA transcription.
• mRNA is translated into viral proteins for capsid assembly.
• The newly synthesized (+) ssDNA is packaged into capsids for virus export.

Group III: dsRNA Viruses

• Carry RNA-dependent RNA polymerase (RdRP) to transcribe dsRNA into (+) ssRNA transcripts (mRNA).
• Host ribosomes translate mRNA into viral proteins.
• RdRP replicates viral RNA, generating (-) strand copies to form dsRNA for new virions.
• dsRNA is packaged into capsids to form progeny viruses.

Group IV: (+) ssRNA Viruses

• The genome functions as mRNA but is not directly translated.
• Host ribosomes translate RdRP, which synthesizes large numbers of (-) ssRNA templates.
• (-) ssRNA serves as a template to make new (+) ssRNA genomes and mRNA.
• mRNA is translated into viral proteins for capsid assembly.
• Some (+) ssRNA is packaged directly as genome material for new viruses.

Group V: (-) ssRNA Viruses

• This is the largest family of RNA viruses; all are enveloped.
• Host cells cannot transcribe (-) ssRNA, so the virus carries RdRP in the virion.
• RdRP synthesizes complementary (+) ssRNA, which serves as mRNA.
• mRNA is translated into viral proteins.
• RdRP also replicates the (-) ssRNA genome for packaging into new virions.
• RNA viruses mutate rapidly because of error-prone replication (e.g., Influenza).
• RNA viruses tend to have small, segmented genomes and high mutation rates due to the instability of single-stranded RNA.

Group VI: (+) ssRNA Retroviruses (e.g., HIV, SIV)

• Reverse transcriptase (RT) copies (+) ssRNA into an RNA-DNA hybrid.
• RNA is degraded, and DNA-dependent DNA polymerase (DdDP) converts the remaining DNA strand into dsDNA.
• dsDNA is integrated into the host genome.
• Host transcription produces new (+) ssRNA, which serves as mRNA and viral genome.
• Progeny viruses are assembled with newly synthesized RNA and proteins.