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174 Cards in this Set

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define virus
a virus is a small, infectious, obligate intracellular parasite, capable of replicating itself in a host cell
List the 5 common properties of viruses
1. obligate intracellular molecular parasites that are small and infectious
2. genome of RNA or DNA
3. Virus genome directs the synthesis of virion components withing a permissive host cell
4. Progeny virus particles are produced by assembly of newly made viral components
5. Progeny virus particles spread infection to new cells
What type of genome do Baculoviridae have?
What do they infect?
DNA virus that infects insects
What is the unifying principle of virus particles
All viruses package their genomes inside a protective particle to facilitate and ensure transfer from one host to the next
What is the unifying principle of viral genomes
A virus's genome contains the information needed to initiate and complete an infectious cycle within a susceptible host.
What is the unifying principle of virus relationship to host
All viruses can establish themselves in a particular host population and thereby ensure their long term survival
Describe the common aspects of the viral life cycle
1. Attachment and entry
2. new synthesis of viral mRNA and proteins
3. genome replication,
4. assembly of new particles
5. release
List the 6 steps of the viral life cycle
1. Attachment
2. Penetration
3. Uncoating
4. Biosynthesis
5. Assembly
6. release
(remember A PU BAR)
What did Edward Jenner do
-British doctor
-vaccinated kid with cowpox
-later challenged kid with smallpox and he didn't get it
What are Koch's postulates
1. the microbe must regularly associated with all organisms with disease but not in healthy animals
2. must be isolated from disease host and grown in pure culture
3. the cultured microbe should cause the disease when introduced into a healthy host
4. same microbe must be reisolated from the inoculated diseased host and identified as the orignial causitive agent
Describe the procedure to develop a one step growth curve
1. inoculate a uniform population of cells with a high dose of virus and then dilute the culture. This ensures that all cells are infected at the same time
2. At time intervals collect a sample of the cells and culture medium
3. measure the concentration of infectious virus both inside the cells and in the medium
What three lines are drawn on a one step growth curve
1. total virus
2. released virus
3. cell associated virus
What is the eclipse period?
The phase of viral infection during which the nucleic acid is uncoated from its protective shell and no infectious virus can be detected inside the cells
Where is infectious virus detected during the eclipse period
nowhere, the virus has been uncoated and synthesis is occurring in the cell but there are no finished particles
What is the latent period
The phase of viral infection during which no extracellular virus can be detected
Where is infectious virus detected between the eclipse and latent period
inside the cell. since the eclipse period is over, there are infectious particles inside the host cell but they have yet to escape
where is infectious virus detected after the latent period
mostly outside of the cell. As cells begin to release virus and die (for a lytic virus) most of the new particles are outside the cell and few are left inside intact cells
How are viruses classified in the classical system
classified by basic properties including genome, symmetry of capsid, presence or absence of an envelope, and dimensions of the virion and capsid
What are the three systems used to classify viruses
1. classical
2. diease based
3. Baltimore
How are viruses classified using the disease based system
classified based on type of disease cause, important for medical and public health, similar sxs does not always mean the viruses are biochemically similar
How are viruses classified using the Baltimore system
classified based on the the nature and polarity of the viral genome, describes how they get to an mRNA
What nucleic acid intermediate is common to all viruses
All viruses must somehow make a +strand mRNA so that the virus can use host translation machinery to make viral protein
define virion
-composed of outer protein shell (capsid) and the interior viral genome
-the virion is the COMPLETE virus particle
This the 3 common properties of virus capsids
-must deliver viral genome to host cell
-must be stable enough to protect the genome but sufficiently unstable to release the genome upon entering the cell
-comprised of repeating protein subunits
why must viruses use repeating proteins in their capsids
the coding capacity of viruses is insufficient to build a particle with unlimited capsid protein types, repeating proteins also allow the virus to self assembly
Describe helical viral capsids
-cylindrical shape with rotational symmetry
-multiple identical subunits
-irregularly shaped proteins around the circumfrence
-virus genome is contained within the hollow center of the capsid
Describe the icosahedral virus capsid
-common among simpler viruses because there is limited space for the genome
-there are two, three, and fivefold axis of symmetry
-usually multiple proteins per face (eg. poliovirus has VP1, VP2, VP3)
Where do Pox viruses replicate?
in the cytoplasm
Why is simply counting virus particles a poor way to detect viruses
not all particles are infectious
What are the advantages of using animal models for detecting viruses? Disadvantages?
advantage-natural infection, can test Koch's postulates, enables study of pathogenesis
disadvantage-expensive, variation among individuals, ethical issues, can always infect w/ human virus
How might virus infectivity be measured in cell culture
1. plaque assays
2. endpoint dilution aka TCID 50%
3. ELISA
What is the difference between primary and continuous cell line cultures
primary- cells derived from the animal, limited life span, normal chromosomes, contact inhibited
continuous- immortalized, indefinite passages, dedifferentiated, aneuploid, tumor cell lines
What are the advantages to using cell lines to study viruses? Disadvantages?
advantage- easy, cheap, molecular studies are facilitated
disadvantage- physiologically relevant?
Describe how plaque assays are used to enumerate viruses
1. infection causes changes in the cell (death/ lysis, protein production, antigenicity, cell fusion, abnormal growth/ morphology)
2. Can use these cytopathic effects to quantify the virus concentation (viral titer)
Perform the assay by serially diluting the virus and infecting a monolayer, incubate cells then stain with a vital dye, number of plaques=> PFU/ml
what is the LD50
the amount of virus need to kill 50% of the cells in culture
Leathal Dose 50%
How can viral proteins be detected
1. radiolabeling
2. immunoprecipitation
3. immunoblotting- western blot
what are the two minimum requirements for viral attachment
1. virion attachment protein or complex
2. cellular receptors and co-receptors
How is virion attachment typical of a receptor-ligand interaction
the reaction is
-saturable
-specific
Virion attachment to the host cell is multivalent. What is the main consequence of this?
The virion is able to make multiple attachment which effectively increases the dissociation constant with each binding. This makes the binding essentially irreversible
Name two viruses that use a single protein for both attachment and fusion
influenza, vesicular stomatitis
Name two viruses that use two separate proteins for attachment and fusion
measles, alphaviruses
Name two viruses that have multiple proteins functioning as a fusion complex
Herpes simplex virus-4 proteins
Poxviruses-8 proteins (at least)
What is the cellular receptor and co-receptor for HIV
CD4= receptor
CCR5/CXCR4= co-receptor
What types of molecules on the host cell can serve as viral receptors
-proteins
-carbohydrates
-glycolipids
-others
Influenza virus uses host sialic acid as a receptor. How does this receptor determine host specificity
Host specificity depends on the type of linkage used in the the sialic acid
humans have alpha 2,6 linkages
birds have alpha 2,3 linkages
pigs have both
Describe the HIV receptor and co-receptors
GP120 on the HIV particle binds CD4 and then the co receptor. this leads to a change in confirmation of GP120 which exposes the fusion peptide. The conformational change is irreversible the co-receptor ensures specificity
Give two examples of how receptor binding by a virus can activate host signaling pathways
1. Cytomegalovirus glycoprotein B activates the JAK/STAT pathway and interferon production
2. Herpes simplex virus binding to certain cells induces IL-6 production, gD is the likely viral protein involved
What are the two basic entry methods used by viruses to get past the host lipid membrane
In both methods, conformational changes in entry proteins expose hydrophobic domains that allow the virus to gain entry
1. Receptor/co-receptor interaction at the plasma membrane triggers a confirmation change (HIV, Herpes)
2. Receptor mediated endocytosis coupled with low pH and acid active protease trigger the confirmation change and expose the hydrophobic region (Herpes simplex virus, influenza)
Where are the three places that a virus may uncoat?
1. Plasma membrane
2. Within an endosome
3. At the nuclear membrane
What is the trigger for the confirmational change in the fusion protein in plasma membrane fusion
receptor/ co-receptor binding
What is the trigger for the confirmational change in the fusion protein in the endocytic route
acidic pH and/or acid sensitive protease
How does Adenovirus, a non enveloped virus, enter the cell
disruption of the vesicle membrane after endosome acidification
How does poliovirus, a non-enveloped virus, enter the cell
Polio forms a pore, the hydrophobic region of the peptide flips out and forms a pore in the lipid bilayer, the nucleic acid is inserted through the pore
Describe how Reovirus enters the host cell
It "melts" through. The particle enters the cell via receptor mediated endocytosis. The low pH in the lysosome activates host proteases that modify the viral capsid (the sigma 3 protein is removed from the u1 protein exposing the hydrophobic domain). A subviral particle called an ISVP penetrates the lysosomal membrane and enters the host cell cytoplasm.
List four methods for viral uncoating
1. low pH of endocytic vesicle de-stabilizes capsid e.g. influenza
2. Proteolytic digestion de-stabilizes capsid
3. Pore formation and release of RNA e.g. Polio
4. DNA viruses inject DNA through nuclear pore
Explain how viral DNA gets into the nucleus
The particle is trafficked through the cytoplasm to the nucleus using NLS signals. The particle binds to a nuclear pore and the 5 fold axis of symmetry opens and the DNA is injected into the nucleus.
Describe how membrane fusion occurs
the hydrophobic fusion peptide is inserted into the target membrane. A conformational change brings the lipid bilayers into approximation and the membranes fuse
Define a productive infection
a productive infection leads to manufacture and release of new infectious virus particles, usually with host cell dying
Define permissive cells
permissive cells enable a productive infection because they permit a complete replicative cycle to occur
Define non-permissive cell
cells that lack some of the functions necessary for complete lytic cycle of infection to take place
Define abortive infection
an infection that is blocked at any step prior to the appearance of infectious virus. The cells can survive an abortive infection
Define chronic infection
any infection where infectious virus is present and can be isolated at any time. Hosts are sources of infection at all times
Define latent infection
an infection that alternates between no infectious virus being detectable and infectious virus being present. Reactivation from latency can result in disease or asymptomatic shedding from the host. Most human DNA viruses can cause latent infection. Latent infection is permanent
In general, what is the purpose of the early genes of a DNA virus
early genes encode regulatory and replication functions
In general, what is the purpose of late genes of a DNA virus
late genes encode structural proteins, proteins needed for assembly, and proteins needed for release
How do small and large DNA viruses differ in terms of early and late genes
small DNA viruses- early genes and proteins encode regulatory and replication proteins
large DNA viruses- immediate early genes encode regulatory proteins, delayed early genes encode replication proteins
How does Adenovirus make sure that the right genes are expressed at the right time
The genome is clustered so that proteins with related functions are transcribed and expressed at the same time
How does HSV (Herpes) make sure that the right genes are expressed at the right time
HSV has a "jumbled" genome with the early and late genes mixed. Each ORF has its own promoter this is in contrast to Adenovirus that has a "clustered" genome
Which DNA viruses do not trick the host cell into starting S phase
Parvoviruses
Why do DNA viruses (except Parvoviruses) need to trick the host cell into starting S phase but stop the process just before host cell DNA replication would begin
This trick puts the host cell in the optimum state to support viral genome replication. All of the necessary host proteins are available.
Which type of DNA virus viral genes are used to trick the host cell into starting S phase
Early or Immediate Early regulatory genes
How can DNA viruses transform host cells into cancerous cells (basic)
DNA viruses trick cells into starting S phase when they should not. If the infection is aborted and the cell survives while expressing these viral proteins, the host cell can begin to replicated in an uncontrolled manner
What is the main feature of the restriction point that regulates a cell's entry into S phase
Rb phosphorylation. Rb must be phosphorylated in order for the cell to enter S phase.
what happens to a cell if unscheduled DNA synthesis occurs
If there is unscheduled DNA synthesis, p53 activates and apotosis occurs
Describe How SV40 LT, HPV 16 or 18 E7 and Ad2 EIA all trick the host cell into starting S phase early
These proteins bind RB and mimic it's phosphorylation. This gives the "ok" for the cell to start S phase. Normally, when Rb is phosphorylated, it can't bind E2F (a transcription factor). This leaves E2F free to bind DNA and increase transcription. If Rb is dephosphorylated it binds E2F and transcription is blocked. The viral proteins bind to Rb and prevent it from binding E2F thus allowing transcription to occur
In general, what causes cell transformation? What happens to transformed cells?
-continued expression of viral proteins that bind pRB and p53 in a cell that survives causes transformation
-transformed cells are immortalized, lose contact inhibition, and can form tumors in animals
Which DNA virus families can cause latent infections
Parvovirus, Polyoma virus, Papilloma virus, Adenovirus, Herpes virus
T/F infectious virus is present during latency
false, infectious virus is not present during latency
What two proteins must DNA viruses interfere with to trigger early S phase
First they must interfere with pRB so that the cell can enter S phase. This interference however causes unscheduled DNA synthesis which would normally trigger the cell to undergo apoptosis. Instead, the virus also attacks the apoptotic pathway by interfering with p53 or another protein.
T/F all transforming viruses cause human cancer
false
T/F RNA viruses use host enzymes to replicate their genomes
false. host cells have no polymerases that can use RNA templates. All RNA viruses must supply their own RNA dependent RNA polymerase (replicase)
What are the two strategies that RNA viruses use to supply their own RNA dependent RNA polymerases
1. Translate the polymerase as the first step after entry + strand viruses
2. Bring the replicase with you as part of the virion structural proteins - strand viruses
Describe the difference between positive and negative sense RNA
positive-the coding strand of RNA, functions like mRNA
negative- the non-coding or antisense strand, virion polymerase makes mRNA from this strand
Describe the replication schme of a positive strand RNA virus
The positive strand can be used right away as mRNA but it is also copied to a negative strand. This negative strand serves as a template for more mRNAs and genomic RNA.
Describe the replication scheme of a negative strand RNA virus
The negative strand is copied by a viral polymerase into a positive strand. The positive strand can be either an mRNA used to make protein or a template for making a copy of the - strand genome
Describe two ways that RNA viruses get their 5' cap
1. The polymerases of some viruses have a 7 methyl guanosine transferase domain that adds the cap
2. Influenza and Bunyaviruses (Lacrosse encephalitis virus) steal the caps from host cell mRNAs
Describe two ways that RNA viruses get poly a tails on their mRNAs
1. some viruses have a long poly U stretch that is copied
2. Some viruses have short poly U stretches that are copied multiple times due to polymerase stuttering to make the long tail
Describe the two ways in which RNA viruses switch between making mRNA and genomic RNA
1. Post-translationally modify the polymerase so that some of it continues to make mRNA and some switches to make genomic RNA
2. The concentration of nuceloprotein (NP) determines what RNAs are made
List 6 methods that RNA viruses use to get around the monocistronic problem (the host cells can translate only the first open reading frame in an mRNA)
1. use separate promoters for each gene
2. use mRNA splicing
3. translate one large protein then cleave it
4. Segment the genome so that each protein is on a separate RNA thus making a separate mRNA
5.Ribosomal frame shifitng
6. IRES used to force ribosome to initiate internally
What is the purpose of an IRES
example- Picornaviruses
IRES= internal ribosome entry site
This is a strategy to solve the monocistronic problem
the site allows the host ribosome to recognize an internal initiation site on viral mRNA and initiate translation
Give an example of a virus that uses IRES's to solve the monocistronic mRNA problem
picornaviruses
Give an example of a virus that uses a polyprotein and cleavage to solve the monocistronic mRNA problem
picornavirus, togaviruses
What two strategies do togaviruses use to solve the monocistronic mRNA problem
1. polyprotein
2. synthesis of subgenomic mRNAs
Give an example of an + strand RNA virus that uses multiple RNAs to solve the monocistronic problem
Coronaviruses
T/F RNA polymerase move along the RNA molecule
false. The RNA dependent RNA polymerases are usually anchored on a cell membrane and the RNA moves through the complex
List three ways that -strand RNA viruses solve the monocistronic problem
1. intragenic spacer that causes the ribosome to stutter formation of the poly A tail the release of mRNA
2.break up the genome (influenza and Reo)
3. Ambisense genome (arena virus)
Describe how Rhadovirus (-strand RNA) genes are expressed
The genome has poly U intergenic spacers. The polymerase initiates at the 3' end of the genome and transcribes genes in order. At the end of each gene the polymerase stutters at the poly U and the re-initiates transcription of the next gene. About 30% of the time, the polymerase fails to reinitiate and a shorter transcript is made. Thus, the genes closer to the begining of the genome are transcribed more often than those at the end.
How does Rhadovirus (-strand RNA) switch between making mRNA or genomic RNA
the level of nuceloprotein controls the switch. High levels of N protein block polyadenylation by inhibiting stuttering and mRNA clevage
How do Paramyxoviruses (Measles, Mumps, RSV) solve the monocistronic problem
RNA editing, a secondary structure forms that makes the ribosome pause leading to reading frame shift. The change occurs at mRNA translation. FYI In contrast Rhabdovirus stuttering affects the polymease and occurs at transcription
hint: Paramyxovirus "mixes" it up, rrrrRhabdovirus stutters
How do influenza (- RNA) viruses solve the monocistronic problem
segmentation of the genome into eight segments which encode 10 proteins, two proteins are generated from spliced mRNAs
How does influenza (-RNA) get its 5'cap
it steals the cap from host mRNA
How do segmented viral genomes lead to genetic diversity and pandemic flu viruses
infection of a single cel with two strains of virus an result in swapping of genomic segments- Antigenic shift
Describe the mechanism that adenovirus used to solve the genome end problem
pTP covalently binds to the first base on the "leading" strand. This first strand is replicated like normal. The "lagging" strand is protected by DNA binding proteins and then circularized using the inverted terminal repeats. This duplex region serves as a primer and allows for continuous replication of the lagging strand
Describe the rolling circle mechanism that Herpes using to solve the genome end problem
The DNA is nicked and a free 3'OH is exposed. The 3' end is lengthened while the growing point rolls around the circle. The 5' end is displaced and forms ss tail. The 5' end is converted to dsDNA using primers just like in normal DNA synthesis. Note that several genomes are synthesized together and then cleaved
How does Poxviridae solve the end genome problem?
The genome has inverted terminal repeats at the end. This allows the genome to form a terminal loop so the polymerase essentially makes a U turn at the end of the genome and moves to the other strand
What is the purpose of the poly U intergenic sequences in the Rhabdovirus (-RNA) gennome
The poly U sequence causes the polymerase to stutter which makes the poly A tail. After stuttering the polymerase can move on and translate the next gene. Sometimes, the polymease falls off of the RNA and a shorter transcript is made
What role does nuceloprotein play in Rhabdovirus (-RNA) gene expression
Nucleoprotein regulates the transition between mRNA and genomic (-) RNA production. When the concentration of nuceloprotein is high, it blocks polymerase stuttering and prevents transcripts from being made, the polymerase is forced to copy the entire genome
What is the purpose of RNA editing in Paramyxovirus
RNA editing allows the virus to make small amounts of protein. The secondary structure of the genomic RNA makes the polymerase pause and leads to a reading frame shift and a different mRNA
What is unique about Reovirus uncoating and replication
The viral core remains intact. mRNA is release from a pore. Completion of replication takes place in the nucelocapsid, the viral polymerase only works inside the nucelocapsid
Describe how RNA viruses recombine
template switching. The polymerase will fall off one template after synthesizing some of the genome and then finish on another template. This can occur between viral genome and host mRNA leading to the incorporation of host transcripts into the virus
How does the rate of mutation compare between RNA and DNA viruses. What explains the difference?
RNA viruses mutate 100-100,000 times more often then DNA viruses. This occurs because DNA polymerases are more accurate than RNA polymerases, RNA polymerases have no proofreading function, and RNA viruses replicate in the cytoplasm where there is no access to host repair functions
List some consequences of the high mutation rate of RNA viruses
1. drug resistance (more than DNA)
2. immune evasion and vaccine failure
3. Development of quasispecies
What are the two strategies that - strand RNA viruses use to make separate mRNAs from the genome
1. Re-initiation at intergenic spaces
2. segmentation of the genome
What family do the retroviruses belong to
retroviridae
Define retrovirus
Single stranded messenger sense (+) RNA viruses with a host cell derived envelope
T/F retroviruses usually kill the host cell as the end of their life cycle
false the host usually lives
How do retroviruses "reverse" the central dogma
They turn RNA into DNA rather than DNA into RNA
Describe the retrovirus life cycle
1. receptor binding and membrane fusion
2. Uncoating and reverse transcription
3. Transport and nuclear entry
4. Integration into host genome
5. Transcription
6. Translation
7. Assembly and release
Describe how the retrovirus genome is organized
On each end of the genome, there is an identical LTR. The first gene is gag which codes for capsid structural proteins. Next is pol which encodes reverse transcriptase and integrase. Finally there is env which encodes the envelope glycoproteins
Describe the structure of retroviral LTRs (long terminal repeat)
The 3' end of the LTR has the U3 region, the middle contains the R region and the 5' end has the U5 region.
The pbs (primer binding site) is located in the 5' LTR on the 3' end of U5
Describe the ribosomal frame shifting that occurs in retroviruses that occurs to regulate the production of the gag and pol proteins
The viruses needs much more gag (capsid structural protein) than pol. The pol gene is in a different reading frame than gag. 90% of the time gag is translated by itself but 10% of the time, the tRNAs slip backward by 1 base and the pol gene is also transcribed. the Gag-Pol polyprotien is then cleaved to make the separate proteins
Describe how retroviral env proteins get expresse
RNA splicing of the full length genomic RNA leads to the transcript that encodes for the env proteins
Where in the host cell does reverse transcription occur?
the cytoplasm
What are the four enzymatic functions of reverse transcriptase
1. RNA directed DNA polymerase
2. DNA directed DNA polymerase
3. RNAse
4. Helicase
Describe how retroviral proviruses are generated
A tRNA binds to the pbs in the 5' LTR. A double stranded DNA molecule is then synthesized from the + sense RNA. The important features are that the RNA is terminally redundant in the LTRs allowing the second DNA strand to begin in the other PBS
How is the retroviral provirus integrated into the host genome
Integration is carried out by viral integrase (pol gene). Integrase produces a small duplication of host DNA and the proviral DNA is inserted into the host genome at random
what type of genomes do retroviruses have?
+ ss RNA
What two important sequences are encoded in retroviral LTRs
1. signals for host RNA pol II mediated transcription of genomic RNA (promoter and termination sequences)
2. Sequences necessary to carry out reverse transcription of genomic RNA (pbs, etc)
From which genetic material does the information for retroviral assembly and release come from.
The provirus. The provirus must be expressed in order to produce these proteins
List 5 features of virus assembly
1. concentration dependent
2. irreversible
3. complex
4. virus structure dependent
5. host cell compartment dependent
What are the three principal strategies used for viral assembly
1. components spontaneously combine to form particles (Noto, SV40)
2. Particles assemble from precursor polyproteins which are modified by proteolytic clevage to form infections virion (polio)
3. Use of scaffold proteins encoded within the viral genome (herpes simplex)
Describe Herpes Virus Assembly
VP22 serves as a scaffold protein and the other capsid proteins assembly around it. VP22 is then removed by a protease and the pieces are excreted through a pore. The genomic DNA is then inserted through the same pore.
What is the purpose of the M1 protein in influenza assembly
M1 is a matrix protein that serves as a chaperone to bring the -RNA to the cell membrane where the capsid is assembling.
What event must occur in the cytoplasm before picornavirus can leave the cell as an infectious particle
maturation cleavage
Describe the role of matrix protein in retroviruses and the maturation cleavage that affects it.
Matrix protein is connected to the capsid protein and the nucelocapsid that is attached to the genome. The matrix protein effectivley drags the genome to the cell membrane where the virus will bud out from the cell. After budding a protease cleaves the matrix from GagPol creating the infectious particle.
What are the advantages of having an envelope? Disadvantages?
-Advantage=serves an an anchor point for virus encoded membrane spanning proteins that function as viral attachment proteins
-Disadvantage= unstable and prone to disruption by agents like detergents, they are less stable than non-enveloped viruses
Why would a single virus particle be insufficient to initiate an infection (give 3 reasons)
1. some of the virions could have mutations in the capsid that interfere w/ binding, penetration, or uncoating
2. Mutations in the genome that result in a later defect (replication, assembly, release)
3. Viruses are often destroyed before reaching a susceptible cell so a large number is needed to establish an infection
(1&2 occur because viral replication is error prone)
Provide two reasons why a virus would grow in one cell line from an organism but not grown in another cell line from the same organism.
1. One cell line may lack required receptors/ co-receptors needed for viral entry
2. There may be differences in permissiveness later on in the viral life cycle (replication enzymes etc) (abortive infection because the virus enters the cell but fails to complete the life cycle)
Describe two ways in which a fusion peptide may be expose
1. receptor /co-receptor engagement
2. low pH or proteases within the endosome
Describe 4 strategies that small DNA viruses use to express a large number of proteins
1. RNA splicing
2. encode genes in multiple reading frames and use ribosomal frame shifting
3. Use transcriptional regulatory regions and promoters withing the coding region of other genes to economize space
4. Express a polyprotein then cleave it to multiple proteins
What is the principle biochemical event or process in the life cycle of a DNA animal virus that is used to distinguish the early phase from the late phase
the beginning of DNA synthesis
What are the general function of immediate early and early gene products in all DNA viruses
regulation and replication
What five events characterize the late phase of replication of a lytic DNA virus
-virus DNA synthesis
-synthesis of structural proteins
-assembly of virus particle
-packaging of viral genome into particles
-shutdown of host cell protein and RNA synthesis
Explain how to do a TCID 50 (endpoint dilution assay) calculation
Add cell to 96 well plate, serially dilute virus sample so that the rows all have the same dilution and the columns serve as replicates (8 dilutions, 12 replicates of each) Find the lowest dilution that still infected 50% of the wells
Describe 4 methods that viruses use to solve the end genome problem and give an example of a virus that uses the strategy
1. have a circular genome- SV40
2. Use a pTP that is covalently attached to the ends of the genome. pTP marks the start site for DNA replication and ensures that replication initiates at the end of the genome template, the other strand has ITRs that circularize it for replication- Adenovirus
3. Use a rolling circle mechanism that makes a concatemeric DNA (multiple genomes linked together) and separate with nuclease during packaging-HSV
4. Use a DNA genome that is cross linked at the ends so it is effectively circular-Poxviridae
Why do many DNA viruses encode factors that inhibit the host cell's Rb?
DNA viruses don't always encode all of the proteins they need to replicate their genome so they need the host to be expressing these. This requires that the host be in S phase. Inhibiting Rb drives the cell into S phase.
Why must all RNA viruses encode an RNA polymerase in their genome
Animal cells have no polymerases that can use RNA templates to synthesize RNA
List four ways that + strand RNA viruses generate multiple viral proteins from their single strand of genomic RNA
1. segmented genome
2. make a polyprotein
3. use a re-initiation mechanism to generate multiple RNAs
4. Use ribosomal frameshifting or stop codon suppression to generate extra proteins from the same mRNA
T/F + strand RNA viruses utilize their own RNA polymerase to replicate their genome so they must package an RNA polymerase in their capsid.
False, + RNA works as mRNA so they can translate an RNA dependent RNA polymerase as son as the virion uncoates in the host cell. - strand RNA viruses must bring their own polymerase
Describe two methods that RNA viruses can use to acquire a 5' cap
1. the viral RNA polymerase has a 7methyl guanosine transferase activity
2. the virus steals a cap from host mRNAs
How does poly U region stuttering in rhabdovirus control the levels of viral mRNA
About 30% of the time, the viral polymerase fails to reinitaite after encountering a poly U region. The levels of each mRNA are thus determined by their order on the genome because it becomes increasingly likely that the polymerase won't make it all the way through the genome.
Describe two ways that RNA viruses regulate the switch between genomic and mRNAs
1. Post translational modification of the viral RNA polymerase
2. Accumulation of a viral protein (rhabdovirus NP) (NP coats the viral genomic RNA and causes the polymerase to proceed down the entire genome rather than transcribe individual genes)
At what step during the retrovirus life cycle does reverse transcription occur? In which cellular compartment does the reaction take place?
Reverse transcription happens immediately after the virus has uncoated and the RNA enters the cytoplasm. The reaction happens in the cytoplasm
How are the pol an env retroviral genes produced from an integrated provirus (considering the use of monocistronic mRNAs)
-ribosomal frameshifting is used for the gag-pol protein
-RNA splicing is used to generate a separate env mRNA
Which region of a retroviral RNA contains the promoter
the U3 region of the 5' LTR
Which RNA virus replicates its genome in the nucleus
influenza
How do influenza genomic RNAs make their way from the nucleus to the plasma membrane to be packaged without being destroyed by cytoplasmic ribonucleases?
Influenza's negative sense genomic RNAs are coated with viral nucleocapsid proteins. The NPs interaction with the M1 matrix protein and the Nuclear Export Protein (NEP) for escort out of the nucleus and to the plasma membrane.
What type of genome?
baculoviridae
DNA
What type of genome?
Poxviridae
DNA
What type of genome?
SV40 (Polyoma)
DNA
What type of genome?
Herpes Simplex (HSV)
DNA
What type of genome?
Parvovirus
DNA
What type of genome?
Adenovirus
DNA
What type of genome?
Rhinovirus
+RNA
What type of genome?
Polio
+RNA
What type of genome?
Togavirus
+ RNA
What type of genome?
Picornavirus
+ RNA
What type of genome?
Coronavirus
+RNA
What type of genome?
Rhabdovirus
- RNA
What type of genome?
Paramyxovirus
-RNA
What type of genome?
Influenza
-RNA
What type of genome?
Bunyavirus
-RNA
What type of genome?
Arena virus
Ambisense
What type of genome?
Reovirus
dsRNA