Transcript What is latency? - Biological Science

LATENCY, LYSOGENY and SYMBIOSIS
LIVING WITH THE HOST
What is latency?
• a period of quiescence (restricted or no replication or
symptoms) that follows acute infection (virus replication
w/wo symptoms that resolve) and has potential for repeat
bouts of reactivation.
• host defenses are not effective (reduced or limited)
• “repression” of productive cycle genes – restricted
gene expression
• virus genome maintained intact
Lysogeny occurs only in bacteria
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Common to all prokaryotes
Reactivate to lytic by UV
All dsDNA viruses
Maintained and reproduce with cell
Integrated or cytoplasmic
Lysogenic conversion - new host phenotype due to
expressed genes
– Superinfection immunity
– Insertional mutagenesis
– Cell wall structure
– Exotoxins
Latency vs Lysogeny
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What cells
What factors required
State of the viral genome
What genes are transcribed
What proteins are expressed
What are the conditions for reactivation
Phage Lambda
• Nutrient rich medium goes
– lytic (lots of hosts)
• Nutrient poor medium goes lysogenic
• Stages in lysogeny
– Establishment
– Maintenance
– Induction
Lambda genome
Lytic cycle
• Transcription from
two promotors
• IE products are N
protein and cro protein
• N is an antiterminator
for readthrough on the
left and right
• Delayed early on right is
needed for DNA
replication (O, P)
• Q product can turn on late
genes
• Still uses host RNA
polymerase
• Same events on Pl and Pr
• Early termination without
N
• N binds to transcript (not
DNA) with host factors
that read through to later
termination signal
Nut - N utilization site
• Forms structure on nascent
RNA
• Binds with host proteins and
RNA-P
• Late genes made from new
promotor Pr’ in presence of Q
Establishment of lysogeny
• Readthrough from Pr makes CII
• Readthrough from Pl makes
CIII
• CIII stabilizes CII from host
protease Hfl
• CII binds to Promotor for
Repressor Establishment (PRE)
and activates CI (repressor)
gene
• CII also activates genes for
integration
pInt
CI binds to operators on left and right
• Or 1-3
• Or 1>Or2>Or3
• Or1-2 block Pr and activate
PRM (promotor for repressor
maintenance)
• Or3 blocks PRM
• Ol blocks Pl
Decision depends on Right side
• If CRO reaches high
concentration before CI it binds
to Or3 and blocks PRM
• At high CRO blocks Pr
Induction results from destruction of CI
• DNA damage stimulates SOS
response
• RecA protease cleaves CI
• CRO beats CI
What evidence suggests that HSV is a
latent virus?
• Can elicit HSV outgrowth by culturing neurons
with appropriate cells.
• In situ hybridization
• PCR
Why is latency a good strategy?
Long term survival and immune escape
Makes for an opportunistic pathogen
HSV Lytic cycle
• Five IE genes - use host enzymes and host and viral
transcriptional activators
– All IE promoters contain a common cis-acting sequence
(TAATGARAT) that reacts with VP16 tegument protein
– VP16 must interact with two cellular proteins, Oct-1
and HCF, to efficiently induce IE promoter activity
• Cell stimulated towards apoptosis but virus stops events
through several IE gene products
HSV establishes latency in terminally
differentiated nondividing sensory neurons.
• Virus infects and replicates in epithelial tissue, enters the
neuron, travels via neuronal flow to cell body (regional
ganglion)
• May have some replication
• DNA is circular episome; returns via neuron (against the
flow) to epithelial surface.
• Effective immune system resolves primary infection and
enhances establishment of latency. Recurrences diminish
with time (Booster affect?)
• Reactivation triggers probably lead to permissive
conditions – some host factor? or reduced immune
response.
• Evidence that transmission occurs without symptoms – is
there true latency? Is there some reactivation at all times?
Latency associated transcript - major latency
product
• 8 kb processed to stable – 2 and
1.5 kb
– nontranslated RNA
• Promotor for LAT
– has neuron specific
elements.
– is antisense to one of the
immediate early proteins,
but can still get latency with
fragment that does not
overlap. Need 5’ 348 bp
• Is neuron IE nonpermissive
cell?
– ICP4 binds and prevents
LAT transcript in lytic cycle
• Latency is thought to be passive – no viral products are
needed to maintain.
• Reactivation may require transient transcription and at
least a few virions produced.This means the signal changes
the transcription factors that are present.
• Levels of virus DNA in neuron is same in LAT+ and LAT• Conclusion: Lat needed for reactivation not establishment
• Recent evidence that LAT – infection leads to increased
neurovirulence (death of neurons)
• Does LAT protect cell from death during latency
establishment or recurrences thus increasing number of
infected neurons and allowing viral reactivation?
LAT protects neuron from apoptosis
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LAT – give “large number of
TUNEL + neurons at 7 days
TUNEL – (terminal
deoxynucleotidyl transferasemediated deoxyuridine TP nick-end
labeling) fluorescent/chromogenic
label bind to ends of DNA so
fragmented DNA gets more label.
A and D = uninfected
B and E = WT HSV
C and F = lat- HSV
Protection from apoptosis inducers
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Infected three different types of cells with a B-gal gene expression plasmid
and LAT+ (APALAT) or LAT- (BABE) in the presence and absence of
apoptosis inducers.
If LAT reduces apoptosis then more cells with inducer will live and thus there
will be more bgal activity than in absence of LAT gene. Compared to control
plasmid with baculovirus antiapoptosis gene(CplAP)
• The three apoptosis inhibitors work at different points in
the pathway
- Protein kinase
- TNF
- Topoisomerase
- Thus LAT blocks something further in the pathway
- Two other HSV genes are antiapoptotic in productive
infections.
Apoptosis - Programmed cell death
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the death receptor-mediated
pathway (Fas or TNF receptor)
DNA damage through
mitochondrial pathway
ability of LAT to interfere with
apoptosis correlates with its ability
to promote spontaneous
reactivation
LAT enhances neuronal survival
because it has antiapoptotic activity
The concept of viral symbiosis
• Parasitoid wasps - use insect hosts
to develop their larvae
• PolyDNAviruses (PDVs) needed
for success
• PDVs produced in wasp ovaries
and injected into insect with eggs
• Viral gene expression in insect products manipulate host immune
defenses
• No viral replication in insect host
• Endogenous “provirus” integrated
in wasp genome and lost ability to
be “independent”
• May go back 70 million years ago
- vertical transmission
First PDV genome sequenced Oct 2004
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570,000 bp
Composed of 30 DNA circles
156 coding sequences (white)
Only 27% genome is coding
42% of coding DNA has no known
homology
Some known genes
– Protein tyrosine phosphatases
(signal transduction?)
– Transcription factor regulators
– Immunosuppressive proteins
Some genes look like modified host
genes
So what makes this a virus?
• Particles in insect
– DNA may have been from
derived from host
– PDV DNA in wasp DNA at
different regions
– May have been able to
encapsidate in viral protein
• May be a virus that transferred
replicative information to wasp
– Lost unneeded functions