General Virology

General Virology VIRUS STRUCTURE

Virion vs virus • Virion is the infectious particle • composed of nucleic acid, protein capsid, +/- envelope • may be extracellular or intracellular • Virus is any stage of infection

Capsid • Functions • Protection of NA • Attachment for naked viruses • Enzyme • Helical vs Icosahedral Symmetry • Tobacco mosaic virus is a ssRNA virus composed of 6000 nucleotides. The capsid is made of 2100 copies of a single protein subunit that contain 158 amino acids.

Icosahedral symmetry • 20 identical equilateral triangles • Structural units on faces to give morphological capsomers

Envelope • Attachment • Entry • Assembly- matrix proteins • Release • Proteins are viral • Lipids are host • If the membrane envelope is destroyed, the virus becomes noninfectious. Why?

Genome - DNA or RNA • strandedness - (single) (double) • linear or circular, partial double stranded circle • Sense: • Positive-sense, Negative-sense, Ambi-sense • Number: • single, segmented

Proteins • structural proteins • non-structural virion proteins • transcriptase, • protease • integrase

WHAT IS A VIRUS? Viruses: acellular organisms Genomes obligately replicate inside host cells using host metabolic machinery to different extents, to form a pool of components which assemble into particles called virions. FA virus differs from a cell in three fundamental ways: i A virus usually has only a single type of nucleic acid serving as its genetic material. This can be single or double stranded DNA or RNA; ii Viruses contain no enzymes for energy metabolism, thus cannot make ATP; iii Viruses do not encode sufficient enzymatic machinery to synthesize their component macromolecules, specifically, no protein synthesis machinery.

ONE STEP GROWTH CURVE • 1939- Ellis and Delbruck: • Infection with a high multiplicity of infection (MOI): ratio of virus to host cell • Simultaneous infection • Single replication cycle • Sample at time intervals by plaque count for plaque-forming units (PFU), • Identification of latent phase • Determination of burst size/viral yield

FViruses cannot be grown on sterile media, but require the presence of specific host cells. Detection

Virology and Human Disease Tuesday, August 12

Viruses • Infectious agents found in virtually all life forms including humans, animals, plants, insects, and bacteria • Virion Structure • genetic material (either DNA or RNA, double or single stranded, linear or circular) • Protein coat (capsid) surrounding genetic material • Rod (helical), polyheldral, or more complex • with or without a lipid envelope • Viruses are not free living • Unable to reproduce themselves outside of a living cell • Transmit their genetic information from one cell to another • Viruses often damage or kill the cells that they infect

Figure 18.1 Size of a virus, a bacterium, and a eukaryotic cell

Figure 18.2 Viral structure Rod-shaped Icosohedral Enveloped Complex

Figure 18.3 A simplified viral reproductive cycle • Obligate intracellular parasites • Rely on the equipment of cell to replicate • Host range = viruses can only infect certain cell types • Dependant on recognition of host cell receptor • Entry, uncoating, replication, release, cell lysis, infection of new cells

Bacterial Viruses • Bacteriophages – phages • Virulent phage • Reproduces only by a lytic life cycle • Lytic – results in the lysis of the host cell • Phage T4 (infects E. coli) • Temperate phage • Reproduce by either lytic or lysogenic life cycle • Lysogenic - replication does not destroy host • Phage λ (infects E. coli)

Figure 18.4 The lytic cycle of phage T4

Figure 18.02x2 Phages

Figure 18.5 The lysogenic and lytic reproductive cycles of phage , a temperate phage

Animal Viruses • Virus Classification • Genome • DNA or RNA (+/- sense) • Size (kb) • Single or double stranded, linear or circular • # segments, sequence • Morphology • Virion size and shape • Plus or minus envelope • Capsid symmetry and structure • Protein • Biological properties • Physical and chemical properties

Figure 18.6 The reproductive cycle of an enveloped virus • Glycoproteins on envelope recognize receptor on host cell • Viral envelope fuses with cell membrane • Genome and capsid enter the cell • Genome is copied • New RNA genomes • mRNA translated into capsid proteins and glycoproteins for envelope • Capsid assembly • Virus buds from cell

Viral Entry: receptors and fusion • Initial attachment – binding to host cell • Viral surface protein recognizes receptor • Carbohydrates • Lipids • Proteins – transmembrane • Entry – virion conformational change in response to receptor or pH • Location of entry • Plasma membrane (neutral pH) • Endosomal membrane (acidic pH) • Type of entry • Fusion – enveloped viruses • Penetration – nonenveloped viruses

Modification of Host Cell Function • Effects on cellular translation • Viruses activate PKR (cellular kinase) which suppresses cellular translation • Effects through receptor binding • Viral receptor binding may mimic the effects of the natural ligand • Induction of cell proliferation • Viruses need replication machinery and induce cells to enter the cell cycle upon infection • Effects on cellular RNA processing • Inhibit cellular transcription • Degrade cellular mRNAs • Alter RNA processing or export

Viral Replication • DNA viruses • Replicated and transcribed similar to host cell • Examples = adenovirus, herpes virus • RNA viruses • Plus stranded • Translated directly (therefore RNA is infectious) • Virally encoded RNA-dependant RNA polymerase (RdRp) synthesizes more genomic RNA and mRNA for proteins • Example = poliovirus • Minus stranded • Must be transcribed by RdRp for replication and transcription of viral mRNA • Enzyme is carried by the virus into the cell during infection • Example = influenza, measles • Retroviruses – HIV

Figure 18.02x1 Adenovirus Virus infects the upper respiratory tract (common cold) and GI tract (diarrhea)

Figure 18.x6 Herpes • 8 herpesviruses infect humans • Human diseases • Oropharyngeal and genital lesions – herpes simplex • Chickenpox – varicella zoster virus • Congenital microcephaly – CMV (growth retardation) • Burkitt’s lymphoma – EBV (childhood tumor) (mononucleosis) • Kaposi’s sarcoma – KSHV

Figure 18.x3 Paralytic Polio • Summer time epidemic disease – summer of 1885 in England • More than 20,000 cases each year in the U.S. • Infection spreads to brain and CNS and replicates in muscle cells, spreading to motor neurons and causing paralysis • Poliovirus was cultured – Robbins, Enders, and Weller received the Nobel Prize in 1954 • First vaccine developed in 1955 by Salk was inactivated virus • Live attenuated vaccine (Sabin) was approved in 1961 and eliminated virus from the Americas

Simpler infectious agents - Prions Figure 18.10 A hypothesis to explain how prions propagate • Infectious proteins – causing degenerative brain diseases • Scrapie in sheep • Mad cow disease in cows • Creutzfeldt-Jakob disease in humans • Prions are misfolded forms of normal proteins in the brain • Prions induce normal proteins to convert to prion form triggering a chain reaction that increases their numbers

What are viruses? • Small obligate intracellular parasites • Virion • Complete virus particle : nucleic acid + protein coat, which may be surrounded by an envelope • It is the form in which the virus moves between cells or hosts • Viral Genome • EITHER RNA or DNA genome surrounded by a protective virus-coded protein coat (Capsid) • Propagation depends on specialized host cells supplying the machinery for replication, metabolism and biosynthesis

The DNA or RNA genome may be : • ss – single stranded or • ds – double stranded • Genomes may be either: • (+) sense: Positive-sense viral RNA is identical to viral mRNA and thus can be immediately translated into protein by the host cell. OR • (-) sense: Negative-sense viral RNA is complementary to mRNA and thus must be converted to positive-sense RNA by an RNA polymerase before translation.

Definitions • Bacteriophage • Virus that infects prokaryotic (bacterial) cells. • Nucleocapsid: • viral nucleic acid + the protein coat that encloses it. • Represents the packaged form of the viral genome.

Nucleic acid Capsid Envelope protein Membrane protein Viral envelope** Nucleocapsid Viral Structure - Overview Spike protein

Viral Structure • Varies in size, shape and symmetry • VIP for classification • 3 types of capsid symmetry: • Cubic (icosahedral) • Has 20 faces, each an equilateral triangle. Eg. adenovirus • Helical • Protein binds around DNA/RNA in a helical fashion eg. Coronavirus • Complex • Is neither cubic nor helical eg. poxvirus

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Viral Structure Figure 1An array of viruses. (a) The helical virus of rabies. (b) The segmented helical virus of influenza. (c) A bacteriophage with an icosahedral head and helical tail. (d) An enveloped icosahedral herpes simplex virus. (e) The unenveloped polio virus. (f) The icosahedral HIV with spikes on its envelope.

Viral Replication • When a virus infects a cell, nucleic acid must be uncoated and gain access to metabolic machinery of cell. • Virus life cycle is characterized by: • attachment • penetration, with entry of nucleic acid into cell • early expression of virus genes (either directly by translation, if virus contains "+" RNA, or indirectly after transcription and then translation) • replication of virus nucleic acid • synthesis of new virion components • packaging and assembly of new virions • exit from cell

Attachment • specific binding of a virion protein (the anti-receptor) to a constituent of the cell surface (the receptor) • e.g. hemagglutinin of influenza virus • some complex viruses (HSV) may have more than one species of anti-receptor molecule • Penetration • energy-dependent step • occurs almost instantaneously after attachment

After the virus attaches to the host cell, it can enter the cell by several mechanisms: • Transfer of the entire viral particle across the cell membrane by endocytosis • Transfer of only the viral genome through the cell membrane • Fusion of the viral envelope with the host cell membrane

Uncoating • at same time as penetration or shortly after • separation of viral nucleic acid (n.a.) from outer structural components • Released as: • free n.a. (picornaviruses) • as nucleocapsid (reoviruses) = may need acidic pH in endosome • viruses only infectious agent for which dissolution of infecting agent obligatory step in replicative pathway • Expression of viral genome and synthesis of viral components

After the viral nucleic acid is released inside the host cell: • The transcription and translation processes of the host cell are redirected for the production of viral proteins and nucleic acids • The different types of nucleic acid genomes are expressed and replicated in several ways: • DNA genomes undergo replication-using processes similar to cellular replication • RNA genomes may be +ssRNA; Can be read directly as an mRNA or reverse transcribed by reverse transcriptase into DNA • RNA genomes may also be -ssRNA; The RNA must first be used as a template to form +mRNAs

Assembly and Release • Components of capsid synthesis directed by late genes • Assembly of enveloped viruses needs interaction with plasma membrane which has been modified • Final stage of infection • Enveloped viruses released gradually by budding or exocytosis • Naked viruses accumulate in cytoplasm and released during lysis

Prions • Prions • Infectious particles that are entirely protein. • No nucleic acid • Highly heat resistant • Animal disease that affects nervous tissue • Affects nervous tissue and results in • Bovine spongiform encepahltits (BSE) “mad cow disease”, • scrapie in sheep • kuru & Creutzfeld-Jakob Disease (CJD) in humans

Vaccination • Edward Jenner developed the first vaccine in 1798 for smallpox Edward Jenner

Microorganisms as the Medical Enemy • the other side of the picture • influenza: 1918-1919 • killed 20 million people Micrograph of Influenza virus

General Virology