Lecture 7 - Virus/host interactions

Question 1

What occurs when there is an evolutionary pressure on viruses by the immune system?

Answer 1

Viruses develop strategies to escape the immune system. Reponse is not uncontrolled, just sufficient to allow viruses sufficient time and resources to replicate and infect new cells/organisms.

Question 2

What happens when their are evolutionary pressures on the immune system by viruseS?

Answer 2

Develops multiple strategies to clear the infection. This is not unconrtolled immunity, just sufficient to clear the infection whiclst avoiding unneccary damage to the host.

Question 3

Why might the balance of host-virus interactions go wrong to result in damage?

Answer 3

• immune system may be compromised
• virus may be introduced to a new environment

Question 4

Upon encoutering a microbe, what is the action of the cell?

Answer 4

1. Clear the microbe or inhibit its replication (the interferon system)
2. Process the microbe and present it to the cells of the adaptive immune system (antigen processing and presentation)
3. Alert and attract cells of the immune system that will help clear the infection (up-regulation of surface receptors, cytokines, chemokines)
4. Kill itself (apoptosis)

Question 5

What is the process of innate immune activation?

Answer 5

1. Cell senses PAMP (pathogen associated molecular pattern) through an innate immune receptor
2. Initiates a signalling cascade through adapter proteins and kinases
3. Leads to the activation of transcription factors (phosphorylation, stablilistation, translocation)
4. Results in an upregulation of:
   
   1. Amplifiers/positive feedback (INFbeta, TNFalpha)
   2. Effectors/antivirulent (ISGs) proteins
   3. Messengers/initiators of adaptive response (cytokines/chemokines)
   4. Negative regulators of innate immunity (phosphotases, ubiquitin ligases, microRNAs)

Question 6

What are the features of cells that do not express the INFbeta receptor?

Answer 6

Severly immune supressed

Question 7

What is the interferon system?

Answer 7

1. Following PRR activation (pattern recognition receptor: TLR, NOD, RIG-I, MDA-5, Lectins) IRF3 and IRF7 induce transcription of IFNbeta IFNalpha (type I interferons)
2. Type I IFNs are translated and secreted and bind the IFNAR (interferon A receptor) forming a positive feedback loop
3. This induces the second wave of interferon stimulated genes (ISGs) through the action of other transcription factors if the IRF and STAT families

Question 8

Give examples of how viruses escape the IFN response?

Answer 8

• In Influenza: NS1 binds dsDNA and sequesters it from being sensed by PRRs.
• In Influenza: NS1 inhibits the catalytic activity of PKR
• In KSHV: viral IRFs inhibt host IRF function (compete)
• The HCV NS3-NS4 protease cleaves sinalling proteins (TRIF and MAVS)
• The HCV NS5A proteins inhibits PKR activiation
• HBX protein from HBV also interacts with MAVS leading to its degradation

Question 9

Why must viruses use multiple strategies to escape the interferon response?

Answer 9

Important for the success of the virus

Makes interaction more robust, the immune system doesn’t just have one strategy to activate the pathway

Question 10

what is the purpose of activation of the IFN response?

Answer 10

Once activated, an array of antiviral genes are transcribed - Interferon Stimulated Genes

Question 11

How is the IFN response regulation?

Answer 11

regulted by STAT1, promoters are downstream of interferon alpha and interferon beta

Question 12

What influenza protein inhibits the IFN response?

Answer 12

Influenza NS1 inhibits the function of several ISGs including RIG-I, PKR (interferon stimulated gene)< OASL (directs antiviral activity)

Question 13

What are the features of HCV as a disease causing agent? and the interaction of interferon stimulted genes in treatment?

Answer 13

• positive strand ssRNA virus (flaviviridae)
• 170 million people infected worldwide
• 70-80% develop chronic infections
• major cause of liver disease and liver cancer (living with hep C accelerates infections)
• IFNalpha used as a first line therapeutic
• 50-60% of infected will not be cured by IFNalpha

Question 14

How does the pre-treatment immune activation status of a patient with HCV affect the interferon response?

Answer 14

Elavated hepatic pre-treatment ISG expression is associated with poor response

Elavated macrophage (Kupffer cells - in liver ) pre-treament ISG expression is assocaited with a good response

Question 15

What are the differences between HBV and HCV infection?

Answer 15

HBV seems to be better at hiding from the immune system than HCV and potentially more susceptible to IFN therapy, becaue the expression of these genes are not correlated with an Hep B infection as it is with HCV, meaning that the virus is not in such a close connection with them, therefore treating with IFN more likely to work.

Mapped genes correlating with virema in infection with HCV and HBV. HCV lots of genes upregulated including interferon stimulated genes (e.g. MX1, IFI44) whereas HBV had none.

Question 16

What are the methods by which viruses can escape the interferon response?

Answer 16

1. Interfere with PRR sensing/suppress PRR expression or activity
2. Interfere with signalling downstream of PRR activation
3. Encode decoy IFN receptors
4. Encode decoy IRFs (vIRFs)
5. Supress activity of interferon stimulated genes (ISGs) i.e. by suppressing the transcriptional programme induced by interferons

Question 17

What are the methods by which viruses can escape apoptosis?

Answer 17

1. Inhibition of the IFN response
2. Inhibition of the TNFR (TRAIL) and Fas death efector cascades (e.g. herpeviral vFLIPs, HIV Nef)
3. Inhibition of the caspase machinery (e.g. viral inhibitors of apoptosis cIAPs, poxvirus Crma)
4. Inhibition of p53 and the apoptosis transcriptional programme (e.g. HTLV1 Tax, HPV E6, SV40 large antigen)
5. Encoding for pro-survival genes e.g. homologues of the anti-apoptotic members of the BCL2 family (e.g. EBV BHRF-1 and KSHV vBCL2)

or

inhibitors of the pro-apoptotic members of the BCL2 family (e.g. HIV Nef)

Question 18

How can the methods used to avoid cellular apoptosis by viruses sometimes lead to cancer?

Answer 18

p53 is a tumour supressor gene.

Examples:

1. EBV: viral oncogene: LMP1, induces Burkitt’s lymphoma, lymphomas, Nasopharyngeal carcinoma
2. HPV: viral oncogene: E6 or E7, induced cervical cancer, head and neck carcinomas
3. KSHV: viral oncogene: vGPCR, induces Kaposi’s sarcoma, lymphomas
4. HTLV: viral oncogene: Tax, induces T-cell leukaemia

Question 19

What is the history of contagious cancer?

Answer 19

• until end of 18th century cancer though to be contagious
• Nooth and Alibert (personal physician to king louis XVIII) first independently challenged this hypothesis: injected themselves with breast cancer cells which resulted in short-lived local inflammatory responses but no tumor establishment
• Peyton rous: used cell free filtrates from tumours to transmit cancer between plymouth rock hens#
• Epstein, Achong, and Barr show a virus (EBV) to cause Burkitt’s lymphoma
• Weiss discovered first transmissable cancer cell - canine transmissible venereal tumour

Question 20

Once a virus escapes the interferon response and apoptotic response what must it then do?

Answer 20

Escape the immune response e.g. by taking the pathway of antigen presentation and inhibit steps in the pathway

Question 21

How to viruses escape antigen presentation?

Answer 21

Interaction of the antigen from the T cell with the APC

1. Downregulate MHC-I and co-stimulatory molecules (e.g. KSHV, K3, K5)
2. Inhibit antigen production by clogging the proteosome (e.g. EBV EBNA1)
3. Inhibit TAP and Tapasin (e.g. CMV US3)
4. Block trafficking of loaded MHC-I through golgi (e.g. HIV Nef)

Question 22

What are the antiviral activities of antibodies?

Answer 22

1. Antibody-dependent cellular toxicity (ADCC) - lysis by NK cells recognising an antibody coated virion/infected cell
2. Completement activation (classical pathway) - phagocytosis following complement activation of antibody coated particles
3. De novo infection inhibition (neutralising antibodies) - antibodies tha tbind and block viral proteins (usually envelope) that are required for viral attachment, binding and entry into host cell

Question 23

How does the virus escape the antibody response?

Answer 23

1. Inhibition of NK cell activation and function (inhibition of ADCC)
2. Inhibition of complement activation
3. MAturation of envelope proteins (influenza HA, HIV gp120 and gp40) - requirement for broadly neutralising antibodies, which is difficult (especially for HIV)
4. Direct antiboies to irrelevent epitoes (HIV gp120) - many times antibodies to immunodominant viral epitopes are not neutralising
   
   • immunodominant proteins are not involved in entry, antibodies bind an inacive conformation of the protein

Question 24

Describe a real life interaction between the virus and the host

Answer 24

• Dramatic increase of Kaposi sarcoma incidence in immunocompromised indiviuals , signifying the beginning of the AIDs pandemic
• AIDs was killing off the immune system, herpes virus switched from not causing the host damage to an infection that killed within 2/3 days
• decrease of disease incidence and severtiy following the introdution of antiretroviral treatment
• Kaposi sarcoma treatment remains the most common cancer in HIV infected indiviuals
• found that cells that don’t have TLR4 are more susceptible to developing the more agressive form of the disease

Question 25

How was it found that people that don’t express TLR4 are more susceptible to the agressive form of kaposi’s sarcoma?

Answer 25

1. First looked at mRNA/protein levels of TLRs during infection and found that TLR4 was the only one downregulated
2. Levels of TLR4 of cells treated with a UV inactived virus compared to a non inactiveated virus. (UV inactivation destroys the genome of the virus, cannot replicate as cannot transcribe its own proteins but can bind to the cell as the coat and envelope not destroyed)
3. Observed a down-regulation of TLR4 even if the virus is inactivated, therefore the binding of the virus results in downregulation. Virus affect the immune status of the cell as soon as it binds by initiating signalling cascades that can lead to the upregulation/downregulation of the appropraite proteins.
4. If the virus is heat inactivated the envelope is also destroyed - get supression but after 72 hours the level of TRL4 goes up. Therefore there must be a second mechanism which relies on viral genome transcripts.
5. LPS bind the TLR4, if it really is downregulated the infected cell doesn’t bind to LPS (non-infeted cells induce TNFalpha). Therefore the receptor of LPS is supressed.
6. Downstream of TLR4 is the interferon pathway and inflammatory cytokines. TLR4 is the only cytokine receptor that can activate both pathways.

Question 26

Why is kaposi’s sarcoma such a problem in africa?

Answer 26

Polymorphism which leads to the more aggresive form of the disease is present in 20-30% of the population in africa complared to 5% elsewhere.

Question 27

How can viral load be measured?

Answer 27

By using PCR

-Applicable for viruses that persist as DNA within cells (dsDNA, retroviruses)

1. Extract genomic DNA from infected cells and perform qPCR or a host gene and a viral gene. This will give the cycle threshold number (Ct) to detect the amplification of each gene (the lower the number the higher the amount of DNA). In the same assay test a standard curve of known copy number of the host and viral genomes
2. Use the host gene to calculate the number of cells. Number of cells = number o fhost genome copies/2 (diploid genome)
3. Use number of viral gene copies to determine the number of viral genome copies
4. Divide the two to get the average viral genome copy number per cell

Viral load = number of viral genome copies/number of cells