WEEK 3: MOLECULAR VIROLOGY AND INFECTION Flashcards
Are viruses ABSOLUTELY DEPENDENT on at least some cellular factors and on cellular metabolic energy?
- YES!
What general terms are viruses known as?
- Obligate intracellular parasites
What are the two phases in a viruses life cycle?
- A VIRION
2. INFECTED CELL
What are virions?
- A SINGLE infectious particle –> what we think of when we say viruses
What are virions inert carriers of ?
- Nucleic acid payload
Which phase of the life cycle do virions participate in?
- The extracellular phase which is a spore
Can virions perform replication or metabolism alone?
- NO!
What process do viruses regulate?
- C, N and P fixation (global cycles)
- They ‘drive’ global cycles
Can a latent virus become active again at another time?
- YES!
Is the virome a major part of the microbiome?
- YES!
Are the vast majority of DNA sequences in human blood viral?
- YES!
- Viral sequences present in ALL tissues
Do ALL viruses make you sick?
- NO
What are 3 examples of q’good’ viruses?
- Polydnaviruses –> Survival of wasp egg in insect host
- Retroviruses –> mammals–> evolution of placenta
- Parvoviruses–> aphids–> needed for development of WINGS
What are 4 viruses that are useful for us?
- Oncolytic virotherapy
- Viral gene therapy
- Reverse transcriptase
- Phage therapy
What does Oncolytic virotherapy involve?
- Viruses with propensity to target/replicate within tumour cells –> Parvivirus
What does viral gene therapy involve?
- Gene therapy using an adenovirus vector
What does reverse transcriptase do and how is it useful?
- Revolutionised molecular biology
- Can generate cDNA from RNA –> useful for PCR, cloning, Sequencing, genomics
What does Phage therapy involve?
- Targeting bacterial diseases and destroying them
e. g. Destruction of MDR bacteria
What are the ‘Historic’ (not used anymore) properties that can classify a microbe as a virus?
- Very small
- But now there are giant viruses so we don’t use it anymore
Do viruses grow when placed in broth?
- NO! THEY DO NOT GROW!!
What is the eclipse period defined as?
- NO new virus present in host cells (immediately when injected, there will be a period of ‘nothing’ happening)
What is the burst size defined as?
- Large number of new viruses produced per infected cell
“Viruses do not grow and divide like bacteria, but are _________, assembled from _____________ and released.”
“Viruses do not grow and divide like bacteria, but are MOLECULAR NANOMACHINES assembled from PREBUILD NEWLY SYNTHESIZED COMPONENTS and released.”
What happens when an extracellular inert particle enters the cell?
- Components dissociate
- Eclipse phase occurs –> NO VIRION detectable
- After some time, protein components made, genome is REPLICATED
- Virions SELF ASSEMBLE from newly synthesized components
Which two components are ALL virions comprised of ?
- Nucleic acid and protein
Can virions have BOTH DNA and RNA?
- NO
- They have to have ONE or the OTHER
What is a nucleocapsid made up of?
- Capsid + nucleid acid
What are many identical units of nucleocapsids called? -
- capsomeres
What is a virion that only has a nuceloplasmid called?
- A naked virus
What is a virion that has a membranous lipid called?
- Enveloped virus
Where will the viral spike/coat proteins that bind to host cell/tissue receptors be found if the virus is NAKED?
-Part of the capsid OR a protein coming off from the capsid
Where will the viral spike/coat proteins that bind to host cell/tissue receptors be found if the virus is ENVELOPED?
-Glycoproteins (bc. inside the envelope)
What are 3 things that the host cell receptors do when binding?
- Recognition of host cell
- Cell entry
- Penetration/uncoating (Destabilisation)
What is the general order from inside to outside of a virion?
- Nucleic acid (DNA or RNA)
- Protein shell /Capsid
- Other layers (Matrix–> HIV) (not always)
- Envelope with glycoprotein (not always)
What is the PRIMARY DEFINING feature of a virus?
- Whether there is an envelope present or not
Apart from the main defining feature of a virus being the envelope, what are the 3 main structures of viruses?
- Spherical
- Helical
- Complex
What are the details of a SPHERICAL virus?
Icosahedral ORDERED SYMMETRY
- Formed from REPEATING UNITS of protein
- Regular polyhedron with 20 TRIANGULAR FACES
What are the details of a HELICAL virus?
Rod shaped coat
- Multiple REPEATING copies of protein coated ONTO GENOME
- Hollow tube–> “spherical staircase”
What are the details of a COMPLEX virus?
Large viruses and a MIXTURE of shapes with NO CONSISTENT SYMMETRY
-Poxyviruses/giant viruses, many bacteriophages, archeal viruses
What predictions can we make from just knowing the structure of a virus?
- Cell entry
- Disassembly, release of genome
- Assembly of virion
- Egress (leaving)
- About stability/transmission Enveloped= usually less environmentally stable
- Naked= More stable and RESISTANT
What do viruses lack the genetic information to encode?
- Machinery….
o To generate metabolic energy, C metabolism
o To generate membranes
o For protein synthesis
Without cells, what are viruses?
- Inanimate, complex organic matter
How are progeny (virions) formed?
-By SELF-ASSEMBLY from newly synthesied components within host cell
Do virions grow and divide?
- NOONONOOO
Do all viruses (even giant viruses) lack a COMPLETE protein synthesis (ribosome) machinery?
- Yes!
Do all viruses have to be able to make mRNA which is translated by host cell ribosomes?
- YES!
What are the 5 basic steps of repliciation of a virus ?
- ATTACHEMNT
- PENETRATION/UNCOATING
- GENE EXPRESSION/REPLICATION
- ASSEMBLY
- RELEASE
What does Attachment involve?
-Enters host organism–> ‘recognizes’ specific target cell or tissue –> ATTACHES TO CORRECT HOST CELL
What does Penentration/uncoating involve?
-Uncoating/release/exposure of genome payload ACCESS TO CELLULAR COMPONENTS
What does Gene expression/replication involve?
-SYNTHESIS of virion components (protein + nucleic acid)
What does Assembly involve?
- SELF- assembly of structural subuints (genome + protein)
- Packaging of nucleic acid, maturation
What does Release involve?
-RELEASE of VIRIONS –> via budding (enveloped virus), lysis (naked virus)
Are virions that are enveloped release via budding or lysis?
- Budding
Are virions that are naked release via budding or lysis?
Lysis
What is a host species tropism?
-Specificity of the virus to PARTICULAR host cells. E.g. HIV virus ONLY INFECTS CD4+ T cells, not other cells based on homologous receptors
What is a cell and tissue tropism?
-Where the particular virus will infect certain tissues based on whether they have the protein on the surface or not.
E.g. Proteins on surface of liver cells rather than blood cells –> Hepatitis virus will go straight through blood until it reaches liver
What do the host cell and tissue receptor determine?
- Host species and tissue/cell trophism
What are 3 things that these receptors do when binding?
- Recognition of host cell
- Cell entry
- Penetration/uncoating (Destabilisation)
Do viruses PARALYZE the host translation machniery?
- YES!
- cell works to synthesise parts for the virus
- Proteins is used for GENOME REPLCIATION and assembly
WHAT DOES TRANSLATION INVOLVE IN VIRUSES INFECTING BACTERIA?
- New genome using viral genome as a template
What two things does the baltimore scheme classify viruses on?
- Nature of the genome nucleic acid
2. Mechanisms to generate the mRNA and to replicate the genome
According to the Baltimore scheme, how many groups of distinct microbes are there?
- 7!
What are the two types of DNA viruses that can exist and what is the polymerase that they use?
- dsDNA snd ssDNA –> use DNA dependent polymerase (RNA for trasncription and DNA for replication) from cellular or viral proteins
What are teh 3 types of RNA viruses and what type of polymerase do they use? -
- dsRNA, (+) ss RNA, (-) ss RNA
- Use RNA dependent RNA polymerases
Can celular life make RNA dependent RNA polyermases?
NO! Viruses must encode and carry the protein
What does positive sense ss RNA mean?
- RNA that goes in the 5’–> 3’ direction and can be translated DIRECTLY by the cell
- Multiple proteins (RDRP, structural) –> nucleid acid alone is INFECTIOUS
What does negative sense ss RNA mean?
- RNA that goes in the 3’–> 5’ direction and cannot be directly translated –> must be converted to mRNA –? virion MUST carry an RNA dependent RNA polymerase protein
- Also must deliver the RDRP protein with the genome
What are the two types of reverse transcribing viruses and what type of polymerase do they use?
- ss RNA RETROVIRUSES
- dsDNA gapped virus
- These sue RNA dependent DNA polymerase (i.e. Reverse transcriptase)
Does cellular life expresse RT?
- NO
- Also does not produce SUFFICIENT RT so virus must encode its OWN
What are the steps for the example of the class I dsDNA virus of Herpes simplex?
- Binds surface receptor Penetration, Destabilisation (nucleocapsid enters cytoplasm)
- DNA enters NUCLEUS CIRCULARISES and associates with HISTONES/ nucleosomes
- VIRAL PROTEIN VP16 is delivered to nucleus to stimulate CELLULAR DNA-dependent RNA polymerase to TRANSCRIBE IE (Immediately early) genes mRNA–> cytoplasm–> translated to protein (normal process) –>nucleus
- IE gene proteins (include transcriptional activators) stimulate EARLY GENES –>VIRAL DNA dependent DNA polymerase –> this REPLICATES the DNA genome –> progeny DNA genome–>Expression of late genes (for STRUCUTRAL PROTEINS) –> Assembly of virus
What is the stepwise gene expression in DNA viruses?
- Immediately early (IE)
- Early (E)
- Late –> Assembly and egress (coming out of the cell)
- There are 2-3 sets of genes that are replicated (generally)
What is an example of a class I dsDNA virus?
Herpes Simplex (HSV)
What is an example of a ssRNA (+) virus?
- Dengue
What is an example of a (-) ss RNA virus?
- Measles
What is guide RNA (gRNA) involved in?
- Replciation of RNA genomes
Do normal cells ahve an RNA-dependent RNA polymerase?
- No
- Viruses must encode the RNA RNA pol.
What do ss RNA (-) viruses have to deliver?
- Must deliver RNA dependent RNA polyemrase (RDRP) protein WITH the genome
What is a retorvirus inserted into the genome via?
- Enzyme called integrase
How does a RNA viruys make mRNA?
- Uses the NORMAL machinery (RNA pol II) to make mRNA –> protein
- mRNA–? genome and assembled with protein
What does a virus need to have?
- Self-assembly to form and release progeny e.g. T4 bacteriophage
How do enveloped viruses egress and where does it occur?
- Budding! Keeps the host cell IN TACT
- Occurs at places where the viral protein (that cell expresses) are embedded –> does this through CELLULAR SIGNALS
How do naked viruses egress?
- LYSIS!
- Via membrane penetrating/destabilising VIROPORINS –> because there is no membrane around virus so will disrupt the cell membrane
What is involved with an ACUTE infection? (and examples)
- Rapid onset, self-limiting
- BRIEF period of symptoms (or asymptomatic)
- E.g. Resolution–> Flu, rhinovirus
- OR death–> Rabies, High % Ebola
What is involved with a LATENT infection and an example ?
- Initial acute but QUIESCENT eith REPEATED reactivation
- Viral gene promoting reproduction not made/ synthesized in SMALL quantities
- Viral genome persists INTACT BUT QUIESCENT productive infection can be initiated later virions
- E.g. Herpes Simplex
What is involved with a PERSISTENT infection and an example?
- LONG TERM
- Primary infection not cleared by adaptive immune response
- Virus particles/genomes continue to be produced/released WITHOUT CELL DEATH non cytopathic continuously or intermittently
- Has VARIOUS mechanisms–> Modulation of the immune response, replication in tissues that don’t have much immune surveillance
- e.g. HIV, Measles virus, Human T lymphotrophic virus
What occurs when the lysogenic reproductive cycle occurs?
- λ Repressor protein EXPRESSED BY LEFT OPERON (lysogenic) INHIBITS THE RIGHT OPERON PROMORTER (lytic)
- Transcription of the LEFT LYSOGENIC OPERON proceeds –> Expression of lysogeny genes and of more λ repressor (+ve feedback loop)
When does the lysogenic cycle enter the lytic cycle and via which pathway?
- After DNA damage
- Via the SOS pathway–> recA produced and DEGRADES the lambda repressor of LYTIC pathway
- Then lytic pathway ACTIVATED
What is lytic also referred to as and why?
- Productive because it does not always hae to lyse the cell (be a lytic virus)
In the example of HSV1 virus what occurs in the transcriptional phase?
-Lytic/latent program and re-entry from latency to productive infection regulated by transcriptional mechanisms
In the example of HSV1 virus what occurs in the initial infection phase?
- Productive –> Epithelial cells, mucosa
In the example of HSV1 virus what occurs in the entry into innervating peripheral nerves phase?
- Trafficking to cell body
In the example of the HSV1 virus, what occurs in the DNA entering the nucleus stage?
- Persists with NO EXPRESSION of IE, E or L genes (‘Lytic genes’)
- Latency transcriptional program (only express latency associated transcript)
What is a summary of the steps in the HSV1 (cold sorevirus) mechanism?
- Transcriptional
- Initial infection
- Entry into innervating peripheral nerves
- DNA enters nucleus
Do neurons replicate/divide so viral DNA can persist without replication in HSV1?
-YES
What is HSV1 reactivated by?
- Stress –> returns to the reinnervarted area
What occurs in Epithelial cells in terms of the HSV virus?
- VP16 delviered to NUCLEUS and has essential cofactor HCF-1
- IE genes then EXPRESSED
- VP16 and ICP0 reduce heterochromatin formation
- Genome then associates with euchromatin!
- SO LYTIC INFECTION
What does HCF-1 and VP16 do together?
- Kick the IE genes off
What occurs in neurons in terms of the HSV1 virus?
- HCF SEQUESTERED into cytoplasm
- VP16 WILL NOT MAKE IE genes so NO TRANSCRIPTIONAL program (LAT expressed instead)
- Genome associates with heterochromatin
- SO LATENT INFECTION
What is the gene associated with heterocheromatin mean?
- It is a Latent infection
What is the gene associated with euchromatin mean?
- It is a lytic infection
What is the HSV genome packaged by?
- nucleosomes
What is heterocvhromatin?
-“ chromosome material of different density from normal (usually greater), in which the activity of the genes is modified or suppressed.”
Are the lytic gene regions active or inactive in HSV genome? (chromatin)
- Inactive
- Chromatin is compacted so the gene is silenced
Is the latency associated transcript (LAT) in HSV1 an open form of chromatin or closed?
- OPEN FORM!
What is the role of LAT (in HSCV1 context)?
- It supresses genome replication and synthesis of IEs
What are the 4 forms of pathogenesis?
- Cell death–> cytopathic effects of a virus (NON enveloped)
- Immunopathology–> Over agressive response
- Immunosuppression
- Oncogenesis
What is an example of a cytolytic virus?
- Polio virus (class V + ss RNA)
- Naked virus
- Cell lysis –> MNs that are NONregenerative
Which two things do viruses use to LYSE cells?
- Viroporins and piconaviruses (VP2 –> important in breaking the cell open)
What are viroporins?
- Hydrophobic proteins that interact with and DISRUPT membranes
What occurs in the FIRST infection of dengue fever?
-1st infection: Activates the B cells, humoral response and memory
What occurs in the second infection of dengue fever?
- 2nd infection by a DIFFERENT serotype (non-protective Igs bing virus, don’t shut the virus down)–> results in attachment/infection of monocytes (Fc receptor) –> results in INFECTION
- Monocytes are NOT normally infected by dengue (only when coated with Ig)
- So dengue virus infected all these cells (monocytes) that it didn’t infect before –> Elevated viral load, cytokine production –> stimulates T cell responses –> plasma leakage and haemorrhaging! ☹
What do protooncogenes normally regulate?
- the cell cycle and there is REGULATION of tumor suppressor and protooncogenes
Which 3 ways can viruses cause cancer?-
- Activation of cellular ONCOGENES
- Expression of their own oncogenes (Retroviruses)
- Inactivation of tumor suppressors (e.g. Polyomaviruses and papillomacviruses)
What happens when viruses cause cancer in terms of the overexpression of cells?
- Early genes are constitutively expressed (always ‘on’) after insertion into host genome–> E7 physically binds and sequesters Rb, E2F released and ACTIVATES S phase
- Viral cause can be treated with vaccination
What are the two types of retroviruses?
- Transducing retroviruses
2. Non-trsansducing retorviruses
What do transducing retroviruses do?
- insert WITHIN a protooncogene
- transcribed TOGETHER
Carries a DOMINANT oncogene
What do non-transducing retroviruses do?
- Inserts NEAR a protooncogene
- Promoters are very STRONG
- So the provirus promoter can activate cellular oncogene in CIS (far away genes)
What is the pathway taken by retroviruses?
- Reverse transcriptase: RNA–>dsDNA
- dsDNA inserts into genome
- Generates transcript–> Used as mRNA (protein) –> When ENOUGH protein transcript is used as genome and packaged
