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Lecture Outline
I. Steps in Virus Entry • Ex. Retroviruses – virological synapse (special
II. Penetration through Cellular Membrane region where viruses are preferentially assembled
III. Intracellular Transport and transmitted in the contact area between virus-
producing cell and neighbor)
Enveloped and non-enveloped viruses have Infection can also spread from one cell to another without
distinct penetration strategies formation of a complete virus particle
• Measles and other enveloped viruses – induce
Enveloped viruses fusion of infected cells with neighboring uninfected
• Use vesicle transport and coupled fusion (“coming cells
together”) and fission (“breaking apart”) of • Results in formation of syncytia (multinucleated
membranes cells)
• viral envelope: transport vessel in passage from o Responsible for cell-cell fusion when
one cell to the next; nucleocapsid is the cargo incorporated into the plasma membrane
of infected cell
Budding
• viral envelope forms around nucleocapsid and • Transmission is limited to cells located next to
other internal components (matrix proteins) at cell each other
membrane In plants, infection spreads directly through
plasmodesmata which connect the cytosolic
• takes place at external plasma membrane
compartments of neighboring cells
• enveloped virion is then released through EC
• These channels traverse the cell wall and provide
space directly or after transport thru secretory
a conduit of transport of viral genomes
pathway
• For some viruses, plasmodesmata are too narrow
•
for passage; virus-encoded “movement proteins”
Fusion of envelope with cell membrane
are expressed to make plasmodesmata wider
• Delivery of nucleocapsid into cytosol of target cell
• takes place at plasma membrane or membranes
of early or late endosomes (membranous
organelles in which the virion is enclosed after
undergoing receptor-mediated endocytosis)
• advantage: viral macromolecules do not need to
pass directly across the hydrophobic interior of any
cellular membrane.
• Ex. Influenza viruses and togaviruses
Non-enveloped viruses
• no lipid membrane
• to exit from infected cell, they rely on cell A variety of cell surface proteins can serve as
death/rupture of cell membranes to release virus specific virus receptors
particles in nucleus or cytoplasm • attachment factors and receptors in PM
• Examples: o include proteins, carbohydrates and
o Adenoviruses – taken up in endosomes lipids
where they proceed to rupture the o have their own cell functions
endosomal membrane, releasing the o highly conserved
virion and other contents in cytosol o unlikely to undergo mutations
o Picornaviruses – undergo conformational o presence or absence of receptors is one
changes during binding to surface of the most important factors to
receptors or after entering endosomes determine the species and cell types that
that allow viral proteins to form can be infected by a given virus
membrane channels to direct genome to • proteoglycans or glycoproteins
cytosol • immunoglobulin family (CD4, PVR, CAR, CD46)
• LDL (low density lipoprotein) receptor family
Some viruses can pass directly from cell to cell
• Multimembrane-spanning transport proteins,
Release of virions from infected cell is not always necessary Integrins,Glycolipids
for transmission of infection
• Ex. Influenza and paramyxoviruses – bind to sialic
• Filopodia – use to infect neighboring cells through acid residues present on glycoproteins and
contact glycolipids
• Infected host cells play active role in transmission, o To avoid from binding to fragments that
with minimal exposure of virus to extracellular do not support infection, they carry
environment neuraminidases which release the
• Also helps avoid recognition of virus by the host virions by cleaving the sialic acid
immune system residues to which they are bound
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• Ex. HIV-1 and Adenovirus – require more than one • Endocytosis occurs via clathrin-coated pits and
type of receptor vesicles
o Primary receptor – sufficient for o Ferry virus particles through network of
attachment and induce conformational
cytoskeletal fibers and transport them
changes
o Secondary receptor (coreceptor) – away from surface and toward the cell
needed to trigger endocytosis or nucleus
penetration o after leaving the cell surface, virions are
o Association with the receptors can occur delivered to early endosomes as other
sequentially because the binding sites for substances are taken up by clathrin-
the coreceptor are exposed only after coated vesicles
interaction of the virion with the primary
o after internalization, virus particle is still
receptor
Receptors interact with viral glycoproteins surface separated from the cytosol by barrier of
protrusions or “canyons” on the surface of the endosomal membrane
virion • Early endosomes – complex organelles
composed of tubular and bulbous membrane
• In enveloped viruses: Specific surface spike elements in peripheral cytoplasm
glycoproteins -serve as binding sites for receptors o Sorting and recycling station for
o Same glycoproteins mediate both incoming fluid, ligands, receptors, and
attachment and membrane fusion; some membrane proteins
exclusively for attachment o Responsible for dissociation of incoming
o Ex. HA domain of trimeric influenza virus ligand-receptor complexes, recycling of
HA protein empty receptors to cell surface, and
• In non-enveloped viruses: receptor-binding sites dispatching non-recycled material to late
are formed either by protrusions or cavities in endosomes and lysosomes to be
capsid surface degraded
o Ex. Adenoviruses: prominent trimeric o Mildly acidic (6.3-5.5), maintained by
fiber attached to each penton base, with membrane-associated vacuolar-type
receptor-binding site located in a knob at ATPases that pump protons from
the tip; in Picornaviruses: indented cytosol into lumen (interior of the
receptor binding site (“deep canyons”) vesicle)
• Interaction between receptor and virus leads to • Incoming molecules sorted for degradation are
changes in both virus particle and cell exposed to successively acidic pH from early to
o Ex. Binding of HIV-1 to CD4 proteins late endosomes, and ultimately to lysosomes
triggers a change in HIV-1 glycoprotein • Late endosomes – have characteristics of
trimers, allowing them to bind to multivesicular bodies
coreceptors in the cell membrane o Contain intralumenal vesicles
• Attachment of adenovirus to integrins o Move by microtubule-mediated transport
activates a signaling cascade inside the cell that into perinuclear region of cell and fuse
temporarily increases endocytic activity and with lysosomes
prepares the cell for the incoming virus
o Activation of signaling pathways – Entry via caveolae and lipid rafts
essential process for entry/replication • Ex. Simian virus 40 (polyomavirus)
o Signaling pathways – promote changes • Small, flask-shaped invaginations in the plasma
in cell architecture and induce membrane are enriched in cholesterol and
micropinocytosis and caveolar/lipid raft- sphingolipids
dependent endocytosis o Invaginations are formed by induction of
membrane curvature caused by tight
Many viruses enter the cell via receptor-mediated multivalent interaction of capsid proteins
endocytosis on the virion with receptor gangliosides
on cell surface
• Majority of viruses depend on endocytosis to be • Caveolae – contain cellular proteins called
first carried into cytoplasmic vacuoles, from which caveolins and cavins
they can then enter the cytosol o Has a role in signaling and uptake of
cholesterol and selected ligands
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membranes closer in the middle of the endosomal membrane -> induce formation of
cluster membrane channel which viral RNA can use to
o Step 5: When later of bound water slip in cytosol -> leaves empty capsid in
molecules that prevents fusion is endosomal lumen -> remains is RNA-free “B-
eliminated in small local spot in the particle”
middle of HA cluster, two bilayers
proceed to fuse. Virions and capsids are transported within the cell
o Hemifusion (outermost bilayer leaflets in vesicles or on microtubules
fuse first) then the inner bilayer halves
Virus particles or capsids that travel in extremely long
o Step 6: narrow aqueous channel of
distances inside cell
internal cavities of the virus and the • Herpes simplex type 1 and rabies virus
cytosol form
o After flickering, fusion channel remains Diffusion of macromolecular assemblies in the cytosol is
permanently open and expands severely limited due to molecular crowding by organelles,
o Nucleocapsids are now exposed and cytoskeleton, and macromolecules
can move to the cytosol
Virions can be transported:
• Class I fusion proteins (retroviruses,
• inside endocytic vesicles (prior to penetration) –
paramyxoviruses, filoviruses)
has the advantage that cell provides all the
o Reveals a trimeric spike standing
systems and signals needed to move the
perpendicular to the viral envelope
incoming vesicle in the direction of nucleus
o Dominated by long central alpha-helical
• or as free nucleocapsids within cytoplasm (after
coiled coils (has 2 distinct
pene)
conformations)
• Class II fusion proteins (togaviruses, molecular motors used include
flaviviruses, rhabdo and hepadna) • dynein – microtubule-dependent motor
o Fusion-activating changes involve major o moves particles along microtubules from
alterations in the interactions between cell periphery in the direction of
folded domains and protein subunits in microtubule-organizing center, close to
oligomers the nucleus
o Undergo only minor conformational o Ex. Herpesvirus and adenovirus capsids
changes at secondary structure use it for retrograde transmission
• dynactin – adaptor complex
Non-enveloped viruses penetrate by membrane
lysis or pore formation
Import of viral genomes into the nucleus
Animal, plant and DNA-containing virus – nucleus as site
Adenoviruses – best-studied example that penetrate by
of replication
acid-activated, virus-dependent endosome lysis
• can establish latent state in nucleus and integrate
• Virion is subjected to changes prior to activation
their genes in host chromosomes
• Fibers dissociate, penton base proteins bind to
• problem: incoming virus must deliver its genome
arg-gly-asp sequences in the integrin and necessary viral proteins to nucleus
(coreceptor) • a number of viruses interact with nuclear
• When virion is exposed to low pH, it converts to
targeting receptors such as importins
lytic form
(karyopherins), and their genomes enter thru
• Endosome membrane ruptures, and virion and
nuclear pore complexes
other contents of endosome is released in the
cytosol Dissociation of nuclear membrane during mitosis
• in retroviruses (except lentivirses)
other non-enveloped viruses pass thru cell membrane
• no means of entering the intact interphase
barriers without lysis
nucleus
• Picornaviruses (polioviruses) undergo pH-
• capsids (“preintegration complexes”) which
independent entry but still depend on endocytosis
contain proviral DNA made by reverse
• Binding to receptor triggers an initial
transcription and other viral proteins must wait in
conformational change -> loosen up capsid wall->
cytoplasm until cell division
makes particles hydrophobic -> these “A-
particles” interact hydrophobically with
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Non-enveloped picornaviruses
• release genome RNA directly in cytosol, leaving
capsid within endosomal vesicle
• virus entry and uncoating are the same event
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