Viruses Flashcards
What is the most common drug hypersensitivity reaction? how does it occur
Morbilliform drug eruption:
amoxicillin treatment for sore throat and fever +
Epstein Barr Virus infection (EBV, mononucleosis)
What is Epstein Barr Virus?
EBV causes Infectious Mononucleosis (IM) also known as glandular fever (kissing disease)
Explain the herpes life cycle
See diagram
What are the three phases of gene expression in herpes
- tegument proteins regulate production of mRNAs and proteins of
IE immediate early genes; they
-protect the virus against innate host immunity
-promote transcription of early genes - production of E early
mRNAs / proteins; they
- are involved in viral replication - production of L late
mRNAs / proteins; they
are involved in virus assembly
Explain how herpes ans EBV illustrate the virus principle
- specific ways to get into the host cells: attachment and virus entry
- Distinguish the virus’ genetic material
Host interactions to establish chronic infection
Expression of viral genes is often in phases - Virus replication, assembly and release
Explain features of Herpes and EBV
dsDNA - Linear in acute, circular in latent phase
120-140kb
120-200nm, envelope, capsid
down-regulate pro-inflammatory responses and MHC-II
can infect B and T cells
widespread infection
Explain points of intervention within the general virus life cycle
- receptor binding (COVID)
- entry (COVID)
- mRNA function- Interferon > RNA breakdown
- DNA/RNA synthesis
- assembly - Protease (COVID)
- release - Neuraminidase (Influenza)
How does INFy orchestrate viral defense?
Type 1 interferon (IFNa - dentrites and IFNb - fibroblasts) produced by infected cells (autocrine and paracrine)
production of >300 gene products and stimulates:
T cells and NK cells produce type II INFy
INFy kills virus infected cells/cancer cells
autocrine - virus replocation in hibition, apoptosis
paracrine - up-regulation of MHC-1 and NK activation
Virus epidemic examples
SARS outbreak 2002/3 - 8,000 cases, 800 deaths nearly 10% fatality
Middle east (MERS): 2012/3 - 1,300 cases, 400 deaths; nearly 40% fatality
COVID19 - 75,000,000 cases, 6,900,000 deaths, 0.9% fatality
what does R0/ the R number mean? give examples
of individuals one sick person will infect
2 = HepC, Ebola, Influenza
4 = SARS, HIV
6 = Pox
10 = Mumps
18 = Measles
Explain the use of Dexamethasone in COVID-19
Corticosteroid
in UK - tested on hospitalised patients with COVID-19
benefits for critically ill patients -reduction in mortality by 1/3 for people on ventilators, 1/5 for people on oxygen
Give examples of different types of approved drugs for virus treatment
Nirmatrelvir and Ritonavir (Paxlovid, protease inhibitor)
Remdesivir (Veklury, nucleotide interfering with viral RNA replication)
Molnupiravir(Lagevrio, same as above)
Sotrovimab (Xevudy) is a “biological” – monoclonal antibody or mAb
Explain features of the drug Paxlovid (CHECK WHAT IT TREATS)
Convenience: orally bioavailable (50% in rats) (95% absorbed from GI)
Efficacy: reduced hospitaliasation and mortality
Mechanism: inhibits the cystine protease Mpro that helps cleave and mature chains of the viral protein
prevents transmission
what are proteases required for in virus infection
virus entry
endosomal release
cleave polyproteins
Describe features of the drug Sotrovimab
Monoclonal antibody
Recognises spike protein of virus
used to treat symptomatic acute covid-19 infection in 12 and above
overcome infection and prevent serious illness
What is a mAB?
Monoclonal antibody:
- Mouse (or other animal) challenged with an antigen
- produces cells that produce specific antibodies
-Cross these cells with an immortal / cancerous cell
- Each cell colony then produces a specific antibody, a clone
Neutralising SARS mAB
Source animal is human
Make a monoclonal antibody, as before
Test effectivity against antigen, here SARS spike protein
Spike receptor binding domain (RBD) targeted mAb neutralise the virus (Graham 2021)
What is a virus factory?
A compartment within an infected cell, made by the virus to protect it from degradation/host defences
Example of a virus that creates a virus factory
Coronaviridae (COVID), Pox, Herpes
Examples of cytoplasmic virus factories
Coronaviridae use double membrane vesicles (DMVs)
in the cytoplasm where the dsRNA is produced
Poxvididae form a viruplasm for viral replication and assembly
Examples of nuclear virus factories
Herpesviridae formnuclear replication compartments(RCs) for viral DNA replication and late gene transcription
Examples of RNA viruses
Paramyxovirus (mumps & measles):
RNA-negative strand (mRNA complement)
linear ssRNA– genome, 15.3 kb
enveloped, size 150 nm
Mumps: infection of the ductal epithelium lading to Parotitis; the mumps virus can cross the brain blood barrier and infect ependymal cells
Paramyxovirus down-regulates innate immunity by interfering with interferon responses
Explain virus classification
How they look: capsid, envelope
Their genome:
This can be either DNA or RNA
Do Viruses violate the central Dogma?
YES
Virus classification by nuclear material (Baltimore classification)
double or single stranded DNA viruses
I- dsDNA: all mechanisms like DNA genomic host cell
II- ssDNA: convert to dsDNA first, then like host cell
double or single stranded RNA viruses
III - dsRNA: RNA used as template for mRNA
IV - ssRNA: a positive strand has the same orientation as mRNA;
requires synthesis of complementary RNA first;
mRNA is then synthesized from the complementary RNA strand
V - ssRNA: a negative strand is complementary to mRNA;
mRNA can be synthesized directly from the genome template
VI: RNA genome but dsDNA intermediate
VII: DNA genome but ssRNA intermediate
Examples of virus in the class 1 classification
dsDNA
use normal cellular mechanisms
Herpes, papilloma, pox
Examples of virus in the class 2 classification
ssDNA
Convert to dsDNA first
e.g. parvo
Examples of virus in the class 3 classification
dsRNA
use normal cellular mechanisms
e.g. reo, picobirna
Examples of virus in the class 4 classification
ssRNA+
must synthesise complementary strand 1st
e.g corona
Examples of virus in class 5 classification
ssRNA-
use the normal cellular mechanism
e.g. paramyxo, filo, orthomyxo
Examples of virus in class 6 classification
ssRNA+, DNA intermediate (retrovirus)
Convert their genome to DNA
e.g. HIV
Examples of viruses in class 7 classification
dsDNA, RNA intermediate
Use an RNA intermediate
e.g. hepadna
estimate of how many viruses on earth
10^31 (2000 known species)
Which Baltimore class of virus is most common
4
Influenza life cycle
Attached to silica acids on the membrane surface and on proteins
Internalised
Sheds Cathrin coated vesicle
Releases ribonucleoproteuin
Makes mRNA that are spliced to make proteins
RW
How does influenza enter cells
attachment
endocytosis
acidification of the endosome
pH induced conformational change of hemagglutinin (surface protein) - exposure of hydrophobic residues
Leads to loss of envelope and release of viral genome
Hemagglutinin features
active, virus form
trimeric membrane protein
bind glycosylated surface proteins on host membrane
Drugs targeting which proteins are being developed to target influenza
drugs against M1 and M2
Two external proteins of influenza
H=Hemagglutinin
glycosylated protein
binds to sialic-acid
N=Neuraminidase
hydrolyses sialic-acid glycosylation
active against Hemagglutinin as well as surface proteins
How is targeting neuraminidase a point of therapeutic intervention
N Allows virus to leave the cell - this is a step of therapeutic intervention e.i inhibition of N enzyme
Examples of drugs that hinder virus release
Tamiflu (Oseltamivir) & Relenza (Zanamivir)
1. neuraminidase inhibitors
2. interfere with the release of the virus from the cell surface
Number of hemagglutinin and neuraminidase types
H = Hemagglutinin, 16 known
N = Neuraminidase, 9 known
Recombination of RNA segments generation of new virus types
Segmental genome of 8 segments
New virus variants are created by changing segments
Explain what classification of virus bacteria, plants and animals are primarily infected by
Bacteria infected by dsSNA viruses (primarily)
Plants infected by SSRNA viruses (primarily)
Animals are infected by all (but ssDNA) viruses
Ebola virus disease (EVD) features
ssRNA virus
Large- not transmitted by air, need contact
Glycoprotein on surface
one of the worlds most virulent, fatal diseases
Explain the genome of ebola
NP = nuclear protein
VP = viral protein
GP = glycoprotein
L = polymerase gene
cleavage by host furin
What are natural hosts for ebola and why is it only found in particular countries?
Climate has to be right
Pteropodidae fruit bats are the natural hosts of Ebola
These bats are only present in the countries infected
bat has a very advanced immune systems - they have the virus but it doesn’t matter
What 3 reactions does reverse transcriptase catalyse
RNA dependent DNA synthesis
RNA degradation
DNA dependent DNA synthesis
What does integration of the HIV genome into the host genome require
Integrase
Explain Integration and retrotransposition of viral DNA into the host genome
- integrase cuts viral DNA
- Attack of viral DNA on target DNA
- Gap filling by DNA repair
- Either side of integrated viral DNA there is short direct repeats of the target DNA sequence
Explain Retroviral-like retrotransposons, common in eukaryotic genomes
LTRs at each end (long terminal repeats)
Explain nonretroviral retrotransposons in eukaryotic genomes
Poly A at 3’ end of RNA transcript; 5’ end is often truncated
a LINE: long interspersed nuclear element (~7000Bp)
no tRNA binding site
What % of our genome are repeat sequences?
50%
Types of transposons
LINEs, SINEs, Retroviral-like element, DNA-only transposon ‘fossils’
Explain DNA-only transposons, common in eukaryotic genomes
Short inverted repeats at each end
Element moves as DNA cut-and-paste mechanism
Features of transposons
inverted repeats required (min 20 bp)
transposase recognises
the repeats
transposase
brings the
ends together
the 3´OH groups can attack the target chromosome
The HIV genome
9700 nt’s long
5’
LTR
Gag – encodes capsid proteins
Pol – encodes reverse transcriptase and integrase
Env – encodes envelope proteins
LTR
3’
Explain the processing of the HIV genome
primary transcript converted into polyprotein
for Pol gene, further poteolytic processing of the polyprotein
Explain the pol gene
a gene encoding a poly-protein
- protease: proteolytic processing of the polyprotein
- integrase: retro-transposition
- reverse transcriptase: RNA intermediate
- ribonuclease H: degrade RNA
what is the gag gene of HIV required for
required to form virus like particles with transcripts
LTR features
repetitive sequences, several 100 bases in length
tRNA binding sites in LTR help reverse transcription
LTR recognised by integrase for retro-transposition
HIV regulatory protein
Gag (capsid), Pol (polymerase) and Env (envelope)
Vif, Vpr, Vpu are regulatory proteins with various functions
Rev regulates nuclear export of the RNA
Nef interferes with protein trafficking
Tat regulates transcription
HIV life cycle
see poster
Explain early HIV synthesis
Splicing of RNA - Rev, Tat and Nef are translated first
happens inside nucleus then transported out into the cytosol
Explain late HIV synthesis
Mature RNA must not be spliced
Rev enters the nucleus
Rev binds to the RRE (rev response element)
Rev protects RNA from splicing
This ensures export of unspliced RNA (from nucleus to cytosol)
Now all viral proteins have been synthesised and there is also unspliced RNA (all of which will be packaged into the virus
Structure of the GAG protein/gene
N-term - MA (pps) - CA(NTD) - CA (CTD) -(pps) SP1 (pps) - NC - (pps) SP2 (pps) - p6 - C-term
3D structure of assembled gag and EM structure
RW
Explain inhibitors that affect the hexamer-pentamer transition of Gag (HIV)
Small molecules that bind to the viral CA protein can be potent inhibitors of HIV infection
Capsid-targeting drugs are predicted to exhibit high barriers to viral resistance, and ongoing work in this area is contributing to an understanding of the molecular biology of HIV uncoating and maturation
Name a caspid inhibitor that shows promise for clinical development
GS-CA1 (discovered in 2017)
Explain HIV cell entry
HIV Env protein
receptor: CD4
secondary receptor for efficient attachment (CCR5)
1st phase of AIDS
secondary receptor for efficient attachment (CXCR4)
2nd phase of AIDS
Examples of viruses that affect immune cells
HIV, EBV, Hep B
Why do so many viruses choose to invade immune cells
blood / immune cells are generally good vessels to travel the body
cells of the immune system end up in lymphoid organs – more cells to infect here
define nucleocapsid
a capsid that encloses the nucleic acid
Icosahedral Capsid features
10-400nm
advantage - can make the capsid from many copies of few proteins
construction principle - 20 triangular faces
Each triangle is made up from (at least) thee proteins
The simplest virus capsid is made up from 60 proteins
What is the T number (in terms of Icosahedral viruses)
Triangulation number
Larger viruses contain more than three proteins per triangle
Multiplier is known as the T number
Known: T = 1,3,4,,7,9,16,25
Explain capsid formation (T=3) and an example of a virus that undergoes this
RW (TBSV)
Jelly-roll motif
RW
Bacterial viruses/bacteriopage conastruction
head: icosahedral capsid
tail: helical
typically dsDNA genome
Example of plant virus and its construction
helical capsid, Ø 15-18 nm,
length 300 nm
single protein subunit
associated with ssRNA+
Explain the structure of viruses that infect bacteria vs plants vs animals
Bacteria - usually head + tail, sometimes enveloped, rarely helical or icosahedral
Plants - usually helical, sometimes icosahedral, sometimes enveloped (less), not head + tail
Animals - usually icosahedral or enveloped, not helical/head + tail
What is Vaccinia vs cowpox vs Variola
Vaccinia: The vaccine virus
Cowpox (CPXV)
Variola: the disease virus (smallpox) - specific to humans
Features of Vaccinia (VACV), Cowpox (CPXV), Variola (VARV)
- enveloped, size 140-260 nm diameter
- linear dsDNA genome, covalently closed, 130-375 kB
- mRNA & protein synthesis in cytosol
E early I immediate L late - replication & maturation in cytosol
- “wrapping” in trans-Golgi membranes
- Poxviridae, virus factories
Vaccinia envelope packaging
RW
Explain the two forms of smallpox
intracellular mature virus (IMV, fusion) and extracellular enveloped virus (EEV, endocytosis)
Both forms are infectious (EEV is more virulent).
Explain poxvirus encoding approx 200 proteins
Early phase:
essential viral proteins transcribed by viral RNA polymerase
uncoating and release of of viral genome into cytoplasm
Immediate phase:
2 hrs post infection triggers genomic DNA replication
Late Phase:
structural proteins
viral assembly
Virus release
direct release: IMV
passage through Golgi: EEV
Smallpox life cycle
RW
Pox: Viral factories
cytoplasmic or perinuclear
recruit specific structures
cell organelles, e.g. mitochondria, Golgi
organise membrane structures: perinuclear replication complexes
exclude host proteins
interfere with signalling
enable viral replication
When was smallpox declared eradicated
1980
Types of vaccines
Live attenuated
Inactivated (killed antigen)
Subunit (purified antigen)
Toxoid (inactivated toxins)
Adenovirus – gene therapy and vaccination
DNA virus
Popular vector for gene therapy in monogenic diseases
Example: CF, cystic fibrosis
Recombinant Adenovirus is being used as COVID19 vaccine
(AZ vaccine, Sputnik 5)
Explain RNA vaccines
delivered in fatty acids LNPs (lipid nanoparticle)
for COVIS - candidates generated in weeks rather than months
Icosahedral virus example
Papovaviridae (italics) - HPV
HPV life cycle
RW
Human papillomavirus HPV features
more than 70 (120) strains
icosahedral capsid
55 nm diameter
dsDNA
8 kB circular genome
encodes:
L1 + L2 capsid proteins, E1 + E2 replication, E4 + E5 assembly / release, E6 + E7 oncogenic proteins
Explain how HPV influences cell cycle regulation
E6 dependent ubiquitination and degradation of p53
E7 deregulates the tumor suppressor pRb
Explain how HPV infections aren’t usually persisrtant, but what happens when they are
- 70 % healed after one year, 90 % healed after two years
- HPV6 and HPV11 cause 90% of all genital warts
persistent infection - risk of cancer
HPV16 and HPV18 cause 70% of all cervical cancers
Viral life cycles
Lytic EBV
- acute
- linear dsDNA
Latent EBV
- genome inactive
- circular dsDNA form
Chronic: Hepatitis B
- long term
- low levels of virus production
Transforming: Pappilloma
altering cell growth
Explain a vaccine against cancer
HPV Vaccination prevents against cancer
HPV6/11 causes 90% of all genital warts
- cervix, vulva, vagina, anus, penis
- transmitted through sexual contact
HPV16/18 cause 70% of all cervical cancers
Cervarix protects against HPV16/18
Gardasil protects against HPV16/18 & HPV6/11
Nobel Prize 2008 to Harald zur Hausen
What protein protects against host immunity in herpes virus and when is it made
Tegument proteins, made in immediate early stage
