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Summary of the last lecture. Virus classification schemes: Classical Recent: ICVT Baltimore Virus Structure: Virion DNA or RNA, virus replication enzymes Capsid, or nucleocapsid Icosahedral and helical Envelope(lipids and carbohydrates from cells) Glycoproteins (Gp, G): spikes

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Summary of the last lecture
Summary of the last lecture

  • Virus classification schemes:

    • Classical

    • Recent: ICVT

    • Baltimore

  • Virus Structure: Virion

    • DNA or RNA, virus replication enzymes

    • Capsid, or nucleocapsid

      • Icosahedral and helical

    • Envelope(lipids and carbohydrates from cells)

    • Glycoproteins (Gp, G): spikes

    • Matrix protein

    • Assessory proteins

  • Virus assembly

    • Spontaneous

    • Protein-protein, protein nucleic acid and protein-lipid interaction (hydrophobic and electrostatic)



Mechanism of packaging genome

  • Direct contact of genome with nucleocapsid or capsid

    proteins: small viruses such as poliovirus contain

    specific proteins that interact with RNA.

  • Specialized viral encoded nucleic acid binding proteins:

    these proteins, called ribonucleoproteins, nucleoproteins

    or core proteins, contain a nucleic acid binding motif in

    the sequence, or recognize a specific packaging signal

    sequence in RNA or DNA. These proteins are highly basic

    which react with negatively charged nucleic acids.

  • Packaging by cellular proteins: unique to papovaviruses, so

    the small viral genome doesn’t need to devote its limited

    genome to packaging.



Cell culture
Cell culture

  • Medium for cell growth

    • -- water, salts, minerals

    • -- amino acids

    • -- glucose

    • -- serum

    • -- antibiotics (penicillin and streptomycin)

    • -- special growth factors (for some cell types)

  • Cell culture

    • -- primary cell culture: prepared freshly from tissue

    • -- cell lines (immortalized cell culture):directly derived from tumor or primary cells transformed with tumor virus or chemical mutagen


Cell culture cont
Cell culture (cont’)

  • CO2 incubator: temperature 37oC, humidified, co2

  • Long term storage: at liquid nitrogen (-140oC). Cells should be frozen quickly in -80oC freezer and thaw rapidly at 37oC waterbath



Cell types
Cell types

Fibroblast-like cells (BHK)

Epithelial-like cells

(Lung carcinoma)


Cytopathic effects cpe
Cytopathic effects (CPE)

uninfected

0.01PFU of virus/cell infected

0.1PFU of virus/cell infected

10 PFU of virus/cell infected


Cpe cont
CPE(cont’)

Intranuclear inclusion body(LM)

Syncytium

Intranuclear inclusion body(EM)


Virus cell interaction
Virus-cell interaction

  • Attachment (reversible)

    -- virus attachment proteins (capsids, or glycoproteins)

    -- receptors or co-receptors (cell surface molecules)

    -- virus tropism ( spectrum of cells that virus can infect)

  • 2. Entry (penetration cross the plasma membrane, irreversible)

    • -- Receptor mediated endocytosis (the most common)

    • -- Cell membrane fusion (usually enveloped virus)

    • -- Translocation: very rare

  • 3. Uncoating( release of nucleic acid from its protein coat)

    • -- uncoating at the plasma membrane (Paramyxovirus)

    • -- uncoating triggered by change in pH (Influenza)

    • -- uncoating in the cytoplama







Virus cell interaction cont
Virus-cell interaction (cont’)

  • 4. Replication and protein synthesis

    • -- In general, early proteins ( viral enzymes, regulatory

    • proteins) are synthesized first, followed by virus

    • genome replication and late proteins (structural

    • proteins) synthesis

  • 5. Assembly and release

    • -- Non-enveloped virus become mature inside the cell

    • and are released by cell lysis

    • -- enveloped viruses mature by budding from the plasma

    • membrane of host cells




One step growth curve
One-step growth curve

Pfu/cell(log)



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