CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
WAGENINGEN UR
Chapter 5: Attachment and entry of viruses into cells
1. Overview of virus replication
2. Animal viruses
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Cell receptors and co-receptors
Virus attachment sites
Attachment of virions to receptors
Entry of animal viruses into cells
Intracellular transport
Genome uncoating
3. Bacteriophages
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
WAGENINGEN UR
Learning outcomes
1. Outline of generalized scheme of virus replication involving
seven steps
2. Describe how animal viruses attach to and enter their host
cells
3. Differentiate between the entry mechanisms of naked and
enveloped animal viruses
4. Describe the roles of cell components in the delivery of
some viral genomes to the nucleus
5. Outline the infection mechanisms of phages
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
At a glance
WAGENINGEN UR
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
WAGENINGEN UR
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Overview of virus replication
1. Attachment of a virion to a cell
2. Entry into the cell
3. Transcription of virus genes into (mRNAs)
4. Translation of virus mRNAs into virus proteins
5. Genome replication
6. Assembly of proteins and genomes into virions
7. Exit of the virions from the cell
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CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Cell receptors and co-receptors
1. Receptors: specific molecules on surface of host cell >< 1 or more
surface proteins (Virion) [highly specific recognition]
2. Co-receptors: second type of cell surface molecules that virus need to
bind to infect the cell.
3. Some cases: binding to receptor causing conformational change in virus
protein >>>> virus bind to co-receptor.
4. Receptors / Co-receptors = glycoproteins with functions: receptor for
chemokines/growth factors; mediating cell-to-cell contact and adhesion.
5. Many receptors = glycoproteins – folded into domains similar to those in
immunoglobulin molecules.
6. Many receptors < regions of plasma membrane – coated on inner
surface = clathrin / caveolin.
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
Cell receptors
WAGENINGEN UR
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Cell receptors and co-receptors
•
Evidence that a cell surface molecule is a virus receptor:
1. Produce of Mono-Ab against cell surface proteins: 1 of Ab blocks
virus binding / infectivity # corresponding Ag = receptor
2. Soluble derivatives of the molecule blocks virus binding / infectivity
3. Normal ligand for the molecule blocks virus binding / infectivity
4. Expression of the gene encoding molecule into virus-resistance cells
>>> makes those cells become susceptible to infection
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Virus attachment sites
1. Each virion has multiple sites that can bind to receptors.
2. Each site = regions of 1 or more protein molecules
3.
In naked viruses:
• on capsid surface (depression (poliovirus) / ridges (foot & mouth
disease virus)
• On specialized structures (fibres / knobs of adenoviruses; spikes of
rotaviruses)
4.
In some enveloped viruses: on surface of glycoproteins
5.
Some virion surface proteins (=virus attachment sites) can bind strongly
to red blood cells of various species > clump (haemagglutination) #
haemagglutinins (influenza viruses – measle virus)
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Attachment of virions to receptors
1. Forces = hydrogen bonds; ionic attractions; van der Waals forces
2. Sugar moieties (present on receptor or virus attachment site) involved in
the force
3. No covalent bonds
4. Initially virion is weakly bound to a cell at only one or few receptors
(reversible attactment – may detach)
5.
If attach remained > more virus attachment sites bind to more receptors
(irreversible attachment)
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Cell receptors and co-receptors
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Entry of animal viruses into cells
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Binding to receptor >>> cross plasma membrane (at cell surface or
endosome membrane)
1.
Endocytosis: used by cells for many functions: nutrient uptake, defence
against pathogens
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Number of endocytotic mechanisms: (clathrin or caveolin – mediated)
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Hi-jack one or more of Endocytic mechanisms to access to host cell.
2.
In some viruses: plasma membrane coated with clathrin (adenovirus) or
caveolin ( simian virus 40) bend around the virions and virions end up in claor-caveolin-coated endosomes > then these proteins are soon lost.
3.
Other viruses: taken up by independent mechanisms of
caveolin.
4.
Endosome may fuse with other vesicles (lysosomes - pH 4.8-5.0 > lowering
pH within the vesicle > further lowered by pumping hydrogen ions across
the membrane (hydrolysis of ATP> ADP) [enveloped viruses]
clathrin and
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Entry of naked viruses
1. Most of naked viruses irreversible attachment of the virion to the cell
surface leads to endocytosis.
2. Plasma membrane “flows” around the virion > more receptors bind >
virion is completely enclosed in membrane # pinches off as an
endosome
3. Endosome contents are part of external environment – virus not yet in
cytoplasm
4. Mechanism of releasing of virus genomes from endosome are not
understood
5. Some naked viruses can deliver their genomes into host cells through
plasma membrane pores.
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
Entry of naked viruses
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CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
Animal viruses
MABI: Plant Virology Lectures
WAGENINGEN UR
Entry of enveloped viruses
* Reversible attachment may lead to irreversible attachment: (lipid bilayers
do not fuse spontaneously / each enveloped virus has a specialized
glycoprotein responsible for membrane fusion:
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Fusion of the virion envelope with the plasma membrane
Fusion of virion envelope with endosome membrane (by Endocytosis )
Fusion proteins are synthesized as part of large protein > cleaved (has at least
2 hydrophobic sequences [ transmembrane seq + fusion seq ] responsible for membrane fusion) link through non-covalent bonds / disulphide bond > become dimer or trimer in virion
envelope.
Fusion seq normally lies hidden >>>become expose (by virus binding to receptor /
or induced by low pH within endosome)
If fusion seq expose at cell surface: infection by fusion with plasma
membrane or endocytosis
If exposure require low pH: only endocytosis
Fusion seq exposed >> insert into target membrane >> pull 2 membrane
together >>> mediate their fusion (release of energy from the fusion
protein)>>> irreversibly change shape
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
Entry of enveloped viruses
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CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
Entry of enveloped viruses
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CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
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Intracellular transport
1.
Virus or their genome deliver to nucleus: using microtubules
2.
RNA viruses replicate in cytoplasm (no requirement enzymes of the nucleus (except
influenza viruses – require cell spicing machinery – genome must be delivered to nucleus)
3.
Retroviruses:
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replicate in nucleus: copy their genome in cytoplasm > wait until mitosis begin #
nuclear envelope temporarily broken down > (virus DNA + associated proteins) enter
nuclear compartment (can only replicate in cells that are dividing)
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Another retrovirus group (lentiviruses including HIV): can be transported into an intact
nucleus – can replicate in non-dividing cells
4.
Most of DNA viruses replicate in nucleus (structural proteins have sequences allow them to
attach to microtubules > cross it); some of them (iridoviruses, poxviruses) replicate in cytoplasm
5.
Motor proteins: move themselves and any cargo along the microtubules
6.
Nuclear envelope: control movement of materials in and out of nucleus
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
Intracellular transport
WAGENINGEN UR
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
Intracellular transport
WAGENINGEN UR
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
WAGENINGEN UR
Microtubules – Nuclear envelope
* Microtubules:
1. Components of the cytoskeleton
2.
Tracts for material (organelles) transportation in the cell
3.
Hollow cylinders: 25 nm diameter / = α and β-tubulin
4.
= plus end (near the plasma membrane) >< minus end (attach to centrosome near
nucleus).
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Nuclear envelope:
1.
composed of 2 membranes (lipid bilayer)
2.
contain 3000-5000 nuclear pores (complex of > 50 protein species)
3.
Pore = basket + 8 filaments (protrude into cytoplasm)
4.
Channel: centre of the pore / molecules (or particles) + carrier proteins (importins,
exportins) / particles <= 25nm can across; larger particles need to shed to be slimmer or
uncoat at nuclear pore
5.
control movement of materials in (ribisomal proteins, histones) and out
mRNAs, tRNAs) of nucleus
(ribosomes,
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Animal viruses
WAGENINGEN UR
Genome uncoating
1. Complete or partial removal of the capsid to release the virus genome
2. The process can take place (depending on the virus):
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•
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At the cell surface (capsid remaining on the exterior surface of the cell)
Within the cytoplasm
At a nuclear pore
Within the nucleus
3. Successful entry of a virion into a cell is not always followed by virus
replication
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host intracellular defences may inactivate infectivity before or after uncoating
Virus genome may initiate a latent infection rather than a complete replication cycle
4. Some cases: uncoated genome survives intact and transcription (or
translation) can begin at the correct location in the cell
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
Bacteriophages
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1. Phages bind specifically to receptors and/or co-receptors on the surface
of cells (or surface of pili, flagella, capsules)
2. Virus attachment sites: tail fibres (phage T4)
3. Attachment is initially reversible > becomes irreversible after attachment
to further receptors and/or co-receptors
4. Infection = entry into the cell of Virus genome (+ few associated
proteins) / capsid, any associated appendages remain at the cell
surface >< animal and plant viruses (entire virion or at least
nucleocapsid enter host cell)
5. Delivery of phage genome require penetration of cell wall / slim layer or
capsule ( lysozyme of virion aid this process)
CANTHO UNIVERSITY
Laboratory
of Virology
BiRDI
MABI: Plant Virology Lectures
WAGENINGEN UR
Learning outcomes
1. Outline of generalized scheme of virus replication involving
seven steps
2. Describe how animal viruses attach to and enter their host
cells
3. Differentiate between the entry mechanisms of naked and
enveloped animal viruses
4. Describe the roles of cell components in the delivery of
some viral genomes to the nucleus
5. Outline the infection mechanisms of phages