Virus life cycle Flashcards
How do helical capsids assemble?
Nucleoprotein (N) assembles around the genome (concerted assembly - linking packaging to assembly of the capsid), driving by N-N and N-genome interactions. Can occur as the genome is synthesised (seen in negative strand RNA viruses). Genome can be on the inside or the outside in a positively charged cleft. RdRp also binds the end of the genome in negative strand RNA viruses
How do icosahedral capsids assemble?
Capsid can be formed around the genome (concerted assembly) or be formed prior to genome insertion (empty shell; capsid often built around a scaffold). Often get formation of intermediate subunit structures. Can be 1 or many protein subunits. Large capsids often need scaffold proteins
Describe the assembly of polyomavirus capsids
Pentameric capsid subunits (5x VP1 + 1x VP2 OR 1x VP3). These form in the cytoplasm. VP1 has an NLS, so the pentamers are transported to the nucleus. Chaperone proteins (e.g. hsp70) bind the C terminal domain ov VP1 and mediate assembly through interactions of the VP1 C terminal domain.
How is the polyomavirus genome packaged?
As a mini chromosome with hyperacetylated histones (for transcription). Packaged via contacts with VP2/VP3. VP1/2/3 bind DNA non-specifically. There is a packaging signal in the genome (ses) which overlaps binding sites for Sp1 transcription factor (GC boxes, which are also important for genome packaging). Sp1 interacts with VP2/3 inhibiting transcription. This interaction may nucleate capsid assembly (concerted assembly)
Where do DNA viruses often do capsid assembly?
Adjacent to PML bodies - parts of the nucleus with functions such as DNA damage repair and antiviral responses
Describe the assembly of adenoviridae
In the nucleus near PML bodies
Hexons and pentons assembly in the cytoplasm with chaperones. The chaperone (protein VI) of hexons has an NLS for nuclear localisation; the NLS for pentons is in the N-terminus of fibre protein. Capsids assemble before the genome is packaged (empty shell) with aid from non-structural scaffolding proteins. Several proteolytic cleavage events are required for maturation
How is the adenovirus genome packaged?
Packaging signal near to the left inverted terminal repeat with 7 copies of a consensus sequence (A repeat). Non-structural virus proteins bind here. The capsid is assembled, and IVa2 is thought to act as an ATPase to drive genome packaging.
Describe the assembly of herpesviridae
Capsids assemble in the nucleus near PML bodies. VP5 (major component) is imported along with a scaffold with an NLS. Capsids assemble around a scaffold (empty shell). Proteolytic cleavage is required for maturation - cleaving the binding site to the scaffold. The scaffold dissociates as the local pH is lowered when DNA is inserted.
How is the genome inserted into herpesvirus capsids?
There is a unique vertex in the capsid of 12 copies of pUL6. It forms a DNA portal for ATPase dependent genome packaging, similar to those found in bacteriophages. There are cis acting packaging signals in the terminal repeats which bind part of the terminase complex. This complex then docks onto the portal and ATP hydrolysis powers a motor to push the DNA inside the capsid. DNA is cleaved between terminal repeats of adjacent genomes (see herpesvirus replication)
How do retroviruses assemble their capsid?
Gag proteins mediate their own transport, self assembly and budding. Concerted assembly. C type retroviruses assemble at the plasma membrane during budding. B the retroviruses assemble at intracellular compartments before transport to the plasma membrane.
How does Gag self assemble and localise in retroviruses?
Myristic acid inserts into the lipid bilayer when Gag interacts with the correct membrane: the MA domain sequesters myristic acid until it interacts with PIP(2) - enriched on the cytoplasmic side of the plasma membrane. Gag then binds the displaced (by myristic acid) unsaturated fatty acid of PIP(2).
How is Gag processed in retroviruses?
In the provirus, Gag hexameters form an immature shell around the RNA with N termini outwards and C termini inwards. When Gag is cleaved, the C termini (containing NC) complexes with the RNA forming the core (fullerene cone or icosahedral). The CA domain of Gag aids formation of pentamers as opposed to hexameters.
What are the packaging signals in retroviruses?
Package 2 copies of the genome. Selectivity due to sequence/structure specific binding of the genome to the Gag protein. The 5’ UTR is important for genome packaging and dimerisation
Describe the interaction between the RNA genome and the nucleocapsid in retroviruses?
The NC domain of Gag has a central basic region (a zinc binding motif) that binds to the packaging signal. It can also bind RNA non-specifically - capsids with no viral RNA in contain cellular RNA (RNA is important to assembly - concerted). After cleavage, NC becomes associated non-specifically with the dimeric genome
How do viruses acquire envelopes?
The viral glycoprotein normally determine the site of viral budding and particle release. If the virus doesn’t bud at the plasma membrane, it must bud into a compartment that is competent for exocytosis so it can be released. The viral protein must accumulate in a certain location (for some viruses this is absolute - no cellular proteins included at all)
What are membrane rafts?
Some lipids (such as sphingolipids and cholesterol) can associate to form a raft. They tend to be small, heterogenous and highly dynamic. Cellular proteins can selectively associate to compartmentalise processes. Some viral proteins also partition into lipid rafts and could be involved in virus budding and concentrating virus proteins
How do alpha viruses bud?
E1/E2 trimers form hexagonal arrays at the plasma membrane. These then interact with the capsid with precise stoichiometry (80:240). These tight interactions induce budding - in theory no cellular proteins required
How does HIV bud?
Myristylated Gag associates with the plasma membrane and forms a patch
This induces membrane curvature and leads to membrane scission by recruiting ESCRT 1 and ALIX-1 which in turn recruits ESCRT 3
Viruses are released and undergo proteolytic cleavage
How do retroviruses bud?
Gag recruits the cellular machinery (components of the ESRCT 1 complex) required for membrane scission. Nothing else is necessary - get VLPs (virus like particles) in the absence of envelope proteins. There is a critical region in Gag for efficient release.
How does influenza A exit the cell?
The virus RNPs bind M1 in the nucleus which then binds nuclear export protein for export through nuclear pores (via an interaction with Crm1). RNPs associate with Rab11 on recycling endosome surfaces where they are transported to the plasma membrane by microtubules. They associate with micro domains containing HA, NA and M2 around the edge of the lipid raft. Budding is driven by M1 polymerisation and scission involving gM2
How do segmented viruses package their genome?
2 models: either random packaging (pack enough segments so enough viruses have a full complement) or recognition of each segment. Current model is that inter segment base pairing creates the packaged complex - evidence of sequences needed for packaging and the ability for mutated sequences to outcompete the wild type
How do flaviviruses replicate and obtain an envelope?
They replicate in deep invaginations of the ER (spherules) that are still connected to the cytoplasm. The structural polyprotein is heavily processed (cleaved). The genome is packaged and the virus travels through the secretory pathway
How do poxviruses assemble and exit the cell?
Within viral factories, a single membrane forms de novo from crescent shaped structures to form the IV with aid from scaffold protein D13. Proteolytic cleavage of core proteins forms the IMV (this is the particle released on cell lysis). Some IMV are wrapped in another double envelope at the golgi with help of F13. Microtubule motors transport IEVs to the cell periphery where the outer membrane fuses to release a double enveloped particle. Actin polymerisation aids the spread to neighbouring cells. Viral proteins are retrieved from the plasma membrane and targeted to viral wrapping sites.
How do non-enveloped viruses leave the cell?
Traditional view was entirely by lysis, though we now know there is some non-lytic release.
How can viruses use autophagy to aid release?
A region of the cytoplasm is enclosed in a double membraned vesicle - the autophagosome, which is normally used to destroy damaged organelles/during starvation. Non enveloped viruses could destroy the internal membrane before exocytic fusion with the plasma membrane for release.
What is cell-free infection?
Virus particles are released form an infected cell, diffuse through extracellular space then bind and penetrate uninfected cells.
What are the advantages and disadvantages of cell-free infection?
Long distance spread within the host, spread between hosts, wide range of available cell types
Inefficient (low moi), virions are accessible to antibodies, complement and other immune system components, is slow
What are some examples of viruses that spread by cell-free infection?
Polio, aden, vaccinia IMVs (cell lysis)
Influenza, norovirus, rotavirus (released at epithelial cell surfaces)
HIV (blood stream)
What is cell associated infection?
Infection occurs at sites of contact between cells. Viruses are released at existing cell-cell junctions/viruses remaining attached to the original cell until cell contacts form/infected cells are induced to form contacts before viruses is released
What are the advantages and disadvantages of cell associated infection?
Rapid and efficient spread to new cells; protection from the environment and immune response; high multiplicity of infection; exploitation of established cell-cell interactions
Restricted range; reliant on cel adhesion mechanisms; transmission to new hosts requires shedding of infected cells (direct bodily fluid contact)
How does HSV-1 spread?
Cell associated infection. Virus is secreted directly to cell contact points (e.g. lateral surfaces in polarised epithelial cells) through the action of the gE/gI complex. The gE complex is thought to interact with cellular mediators of secretion at the cytoplasmic domain and the extracellular domain is thought to be a receptor. The virus remains attached to the cell via interaction with surface proteins such as heparin sulphate proteoglycans.
What do pUL7 and pUL51 do in the secretion of HSV-1?
Tegument proteins pUL7 and pUL51 are also important for HSV-1 secretion. They are likely to interact with envelop and other tegument proteins. They co-localise to focal adhesions which are important for maintaining cell contacts - they stabilise the cell’s morphology and prevent rounding up and detachment. gE/gI complex then directs virions to stable cell-adhesion sites for secretion
How does HIV spread?
Cell-free infection - inefficient
Cell-to-cell contacts for efficient spread
How does HTLV-1 spread?
Human T-cell lymphotropic virus (causes leukaemia). Almost all spread by cell-associated infection through cell containing bodily fluids such as blood, semen and breast milk. Interactions similar to immunological synapses aid spread from uninfected to infected T cell
What are virological synapses?
Receptor containing adhesive junctions that are the initial interaction between uninfected and infected cell. The virus envelope protein stabilises the interaction. Virus envelope proteins and receptors are polarised to contact sites which is then stabilised by cell adhesion molecules (e.g. ICAM-1). Receptor molecule clustering is stabilised by the actin cytoskeleton. The MTOC is polarised towards the contact site. The virions are trafficked to the contact site where they assemble and bud and enter the naive cell.
What is virus surfing?
A method of retrovirus cell-associated spread. Infected cells anchor filopodia which then transport virus to the cell body.
What is extracellular matrix-bound virion spread?
Method of retrovirus cell-associated spread. HTLV-1 can remain attached to the extracellular matrix before interacting with another cell. Virion clusters are delivered to contact points when they interact with a target cell
What is trans-infection spread?
Method of retrovirus cell-associated spread. Dendritic cells can capture cell free virions. Infectious virions can be delivered to target T cells before infection/replication in dendritic cells
