Herpes virus latency Flashcards Preview

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Flashcards in Herpes virus latency Deck (73):

Latency general

evolutionary advantage with a small population size. Due to co-evolution with ancesters.


a herpesviruses.



B herpesviruses

CMV, HHV6 and 7.


y herpesviruses

EBV and HHV 8.


Herpesviruses lifecycle general

Primary infection of mucosal or epithelial cells produces virions. Latency in non-permissive cells maintains population. Stimulation makes these permissive for reactivation, and make virions which infect epithelial or mucosal cells for acute infections.


Stimulation for reactivation HSV

Trauma, UV, hormonal.
Leads to transport by kinesins to periphery.


Mouse models and HSV latency

Mouse models do not get naturally infectedd, or reactivate.


To cause latency...

block IE expression.


HSV IE gene expression

IE promotor = TAATGARAT
α/IE gene promoters bind Oct1. VP16 binds HCF. VP16-HCF complex promotes formation of preinitiation complex.
VP16 may also act by decrease H3 on gene promoters.


VP16 factor leads to expression of ...

many transactivators, including ICP0 and ICP4 which interact to drive early and late gene expression.


Describe key features of the replication cycle of HSV.

Binding, delivery to nucleus, transcription, replication, assembly, egress.


HSV routes of entry.

1) Binding, membrane fusion, delivery of capsid to cytoplasm.
2) Binding, endocytosis, membrane fusion, delivery of capsid to cytoplasm.


HSV: reversible primary attachment

gC and gB bind cell surface glycosaminoglycans (GAGs)


HSV: specific irreversible attachment

gD to nectins, HVEM or 3-O-heparan sulphates.


HSV membrane fusion requisite proteins

gB, gD, gH and gL. gD = specific binding. gH has fusion peptide.


HSV α genes expression - timing.

2-4 hours post infection.


HSV α genes expression - requirements

No prior viral protein synthesis. Cellular factors provide activation for basal levels. VP16 important for efficient transcription.


HSV α genes expression - downregulation

ICP4 self-represses. B gene products downregulate α gene expression.


Function of HSV α genes

5 out of 6 stimulate β gene expression.


Which gene products block cellular silencing machinery in HSV?

ICP0, Us3 and Us3.5


Role of ICP4

ICP4 is a sequence specific repressor and can be a sequence independent transcription factor.


HSV β gene expression - timing

4-8 hours post infection. Timing dependent on organisation and context of promoter elements.


HSV β gene - classification

two classes according to timing, but no rigid criteria.


HSV β gene expression - control

ICP4: interacts with basal TFs to promote pre-initiation complexes. Later groups may require ICP27.


HSV γ gene expression - control

Requires DNA replication (leaky late and strict late). Cis-acting alteration in DNA template changes protein interactions.


HSV γ gene expression - cellular proteins and etc.

Cellular proteins such as SP1 are used. Gene promoter elements contain TATA box.


HSV genomic replication - first step

Binding of UL9 and ICP8 to origin sequences recruits complex. Bidirectional.


HSV genomic replication - type of replication.

Initially θ replication, then rolling circle replication (concatemers).


HSV latency - preventing α genes expression

1) Lack of nuclear host factors - HCF in cytoplasm, limiting amounts of Oct1.
2) Insufficient transport of VP16 to the nucleus.
3) Hormonally regulated repression of gene transcription.
4) LAT mediated repression of gene transcription.
5) Neuron-specific host miRNA targets HSV-1 ICP0 expression promoting latency.


HSV: LATs general

All genes except LAT gene silenced by heterochromatin. Expressed from neuron specific promoter.
Spliced to give several LATs.
act via miRNA or siRNA


HSV reactivation sequence

Stress at periphery --> ?transduction signal? --> ICP0 promoter activation --> ICP0 production --> global derepression and reactivation --> expression of a genes --> expression of B genes.


Latent HSV DNA

DNA lacks free ends
Contextual analyses have revealed a variability in copy number. 10 -
Latent DNA has a nucleosomal organisation.


Histone modifications in HSV latency

Latent DNA has nucleosomal organisation.
Chromatin immunoprecipitation assays how that the LAT promoter is associated with acetylated histone H3, whilst DNA pol gene is hypoacetylated (inactive).
In some cultured neurones, histone deacetylatase inhibitors activate viral gene expression.


Role of VP16 in preventing repression of the genome (latency)

Recruits proteins for demethylation and acetylation.


Role of VP16 in preventing repression of the genome - demethylation

VP16 binds HCF, recruiting to IE promoters.
HCF recruits lysine specific demethylase (LSD).
LSD prevents H3K9 methylation.
H3K9 methylation is silencing.


Role of VP16 in preventing repression of the genome - acetylation

Also VP16/HCF appears to recruit CBP/p300 and cause acetylation of histones associated with IE promoters.


Importance of LATs in latency

LAT negative mutants increase IE gene expression.
Cell lines expressing major LAT are fairly non-permissive for lytic infection.


Activity of LATs in latency

Reduce viral transcripts, protect from apoptosis.
Possibly play a role in maintaining repressed chromatin state (Lat negative mutants have latent genomes enriched in active histon mark).


Role of VP16 in reactivation of HSV.

VP16 usually just a late protein, so people didn't think had a major role.
But has unique regulatory activity in neurons and can be activated in latently infected cells subjected to stress.


EBV latency cells

Infects B cells. These go to germinal centre, undergo positive selection and either become cells making antibodies or memory cells - these latter are the target latent reservoir.


EBV latency

Active maintenance of episome required.
Epigenetic control of promoter (as HCMV and HSV).
Different latency expression profiles.


EBV latency - maintenance of the episome.

Replication by host machinery including DNA pol, when host is dividing.


EBV latency - epigenetic control

Nucleosome positioning
Histone modification
DNA methylation


EBV latency - epigenetic control, nucleosome positioning.

Similar nucleosomal pattern to host.


EBV latency - epigenetic control, histone modification.

Z promoter. Acetylation, phosphorylation or specific trimethylation can cause reactivation.
Highly complex: at a single residue several different groups could be added, all with different effects.


EBV latency - epigenetic control, DNA methylation.

Cytosine methylation.
Depends on complex interplay between 3 DNA methyltransferases.


EBV latency - epigenetic control, DNA methylation, cytosine methylation.

Method to silence immunogenic genes; cytosine methylation within CpG dinucleotides in promoters esp effective.


EBV latency - epigenetic control, DNA methylation, irreversible.

 Suppression by methylation can only be reversed with de novo DNA replication, therefore CpG methylation at the Z promoter, important for reactivation, is low, so that CpG methylation is not a hinderance to reactivation.


EBV latency transcription

List latency associated genes.
3 potential transcriptional programs.
Cascade in acquiring latency.


EBV latency associated genes

6 nuclear antigens (EBNA proteins)
3 membrane proteins (LMP)
A group of complex spliced transcripts
2 small non-polyadenylated transcripts EBER1 and 2.


Transcriptional programs in latency.

Latency I
Latency II
Latency III
Discuss latency promoters


Latency I in EBV

EBNA1 transcribed under Qp. Wp and Cp are silent.
EBNA-1 has major role.


Latency II EBV

EBNA1 and LMP-1 are transcribed.


Latency III EBV

Usually requires immunosuppression. All latency promoters are active.


Latency promoters EBV

6 nuclear antigens are driven by Cp, Wp and Qp. In most latently infected cells, Cp and Wp are silent, and only Qp is active.


Transcriptional program establishing latency

Starts with latency III profile, driving proliferation. Initially Wp active, activates Cp, gene products of this silence Wp.
Cp slowly acquires methylation, as do LMP1/2 promoters. Progressive silencing results in latency I program.


Latency I in EBV. Role of EBNA1.

EBNA-1 binds to origin of replication, is vital to maintain genome copy number in dividing B cells. Most latently infected cells express this program, but some do not. We do not know how they make the choice.


EBNA proteins immunogenicity

Immunogenic, targeted by CTL, counteracting driving of proliferation by virus.


Reactivation - general

A dangerous step as immune surveillence can kill cell.


EBV reactivation factors

Local trauma or systemic stress
Inhibition of mTOR kinase activity
Hormonal changes
Stimulation of B cells by a different infection.


EBV reactivation factors - mTOR kinase.

Inhibition of mTOR kinase activity due to transient interruption of protein synthesis or hypoxia. Possibly this is how local trauma works. Virus prepares to leave the sinking ship.


EBV reactivation factors - hormonal changes

CD8 T cells contribute to latency.


Process of EBV reactivation

BCR signalling, linearization of genome, production of lytic gene products.


Process of EBV reactivation - BCR signalling

Leads to BZLF1 then BRLF1 gnee expression, under control of Zp promoter. Usually has low activity, but can be enhnaced. Affected by cellular TFs present.


Actions of BZLF1 and BRLF1.

BZLF1 - induces expression of IE genes, recruits HATs. Appears to bind some CpG methylated motifs better than unmethylated ones, which explains the activation of lytic replication when the genome is heavily silenced by DNA methylation.
BRLF1 is also a transcriptional activator.


EBV: neoplasias and latency.

Which latency programs for which neoplasias?
Role of EBV episome and latency gene products in neoplasias.
LMP in neoplasias.


EBV. Which latency programs for which neoplasias?

Most are latency I or II. Leiomyosarcomas in HIV and post-transplant lymphoproliferative disease (PTLD) may be latency III. Removal of immunosuppression reverses the latter.


Role of EBV episome and latency gene products in neoplasias.

Affect epigenetic state, plus TFs have effect.
o Latent infection has been shown to repress host tumour suppressor genes via increased DNA methylation among other mechanisms.
o EBNA1 appears to produce telomere dysfunction, possibly by altering host chromatin structures.


EBV. LMP in neoplasias

Highly pleiotropic, function depending on context. Can drive growth, promote metastasis, apoptotic resistance and immunomodulation.


HSV entry into neurons.

entry by pH-independent membrane fusion
Capsid then transported along microtubules to nucleus (uses tegument proteins US3, UL36 and UL37 = retrograde transport complex


HSV latency not static

Persistant cell mediated immune response found to occur in trigeminal ganglia during latency in both mice and humans —> CD8+ cells produce IFNγ and granzyme B to decrease IE gene expression



Spliced into 4 distinct miRNAs
miR-H2 targets ICP0
miR-H3 targets ICP35.5
miR-H4 targets ICP34.5
miR-H6 targets ICP4


ICP0 in HSV reactivation

ICP0 seems to be key —> mutants reactivate with lower efficiency in mouse models and become latent when used to infect normally permissive cells
acts as E3 ubiquitin ligase —> mediates proteosomal degradation of cellular proteins —> disrupts HDAC function to alleviate histone repression
also involved in IFN antagonism (via inhibition of IRF3 and 7 inhibition)
could be that ICP0 activated (has TF binding sites for Oct1, VP16 and HCF) and then removes global depression by histone acetylation —> reactivation