In response to your first question, it is very clear that it would
stimulate the immune system located
inside the neurons.

I will try to summarize these things, hopefully today.
For now...

Key works includes: &TIF, &0, ICP0, immune response, herpes simplex

http://www.virologyj.com/content/1/1/5
Persistent expression of chemokine and chemokine receptor RNAs at
primary and latent sites of herpes simplex virus 1 infection.

Cook WJ, Kramer MF, Walker RM, Burwell TJ, Holman HA, Coen DM, Knipe
DM.

Department of Microbiology and Molecular Genetics, Harvard Medical
School, Boston, MA 02115, USA. david_knipe@hms.harvard.edu.

Inflammatory cytokines and infiltrating T cells are readily detected in
herpes simplex virus (HSV) infected mouse cornea and trigeminal ganglia
(TG) during the acute phase of infection, and certain cytokines
continue to be expressed at lower levels in infected TG during the
subsequent latent phase. Recent results have shown that HSV infection
activates Toll-like receptor signaling. Thus, we hypothesized that
chemokines may be broadly expressed at both primary sites and latent
sites of HSV infection for prolonged periods of time. Real-time reverse
transcriptase-polymrease chain reaction (RT-PCR) to quantify expression
levels of transcripts encoding chemokines and their receptors in cornea
and TG following corneal infection. RNAs encoding the inflammatory-type
chemokine receptors CCR1, CCR2, CCR5, and CXCR3, which are highly
expressed on activated T cells, macrophages and most immature dendritic
cells (DC), and the more broadly expressed CCR7, were highly expressed
and strongly induced in infected cornea and TG at 3 and 10 days
postinfection (dpi). Elevated levels of these RNAs persisted in both
cornea and TG during the latent phase at 30 dpi. RNAs for the broadly
expressed CXCR4 receptor was induced at 30 dpi but less so at 3 and 10
dpi in both cornea and TG. Transcripts for CCR3 and CCR6, receptors
that are not highly expressed on activated T cells or macrophages, also
appeared to be induced during acute and latent phases; however, their
very low expression levels were near the limit of our detection. RNAs
encoding the CCR1 and CCR5 chemokine ligands MIP-1alpha, MIP-1beta and
RANTES, and the CCR2 ligand MCP-1 were also strongly induced and
persisted in cornea and TG during the latent phase. These and other
recent results argue that HSV antigens or DNA can stimulate expression
of chemokines, perhaps through activation of Toll-like receptors, for
long periods of time at both primary and latent sites of HSV infection.
These chemokines recruit activated T cells and other immune cells,
including DC, that express chemokine receptors to primary and secondary
sites of infection. Prolonged activation of chemokine expression could
provide mechanistic explanations for certain aspects of HSV biology and
pathogenesis.

Introduction

Abstract
Introduction
Results
Discussion
Materials and Methods
Competing interests
Authors' Contributions
Acknowledgments
References

Tables

Table 1
Expression of Chemokine Receptors, Chemokines and Cytokines in
Leukocyte Populations

Acute viral infections are usually cleared from the primary site of
infection by the host immune response [1], but some viruses can persist
at other sites in a latent form. Herpes simplex virus (HSV), for
example, causes a primary infection at a mucosal site, which is cleared
within 710 days by the host immune response. HSV, nevertheless,
enters sensory neurons and establishes a latent infection within those
cells. In a mouse corneal model of HSV-1 infection, infectious virus is
detected in corneal secretions and tissue for approximately 7 days [2].
Similarly, infectious virus is detected in trigeminal ganglion (TG)
tissue for up to approximately 10 days [2]. Latent infection is
established by 30 days postinfection (dpi) because no infectious virus
can be detected in homogenates of TG tissue at that time. HSV DNA,
however, is readily detected in latently infected TG for at least 150
dpi [3-5]. Viral gene expression is greatly attenuated during latent
infection because the only abundant viral gene product detected is the
latency-associated transcript or LAT [6]. Nevertheless, low levels of
lytic transcripts can be detected in ganglia latently infected with HSV
[5]. Evidence of viral protein expression is provided by the continued
T cell infiltration [7,8], elevated levels of interferon  (IFN-)
and TNF- transcripts and numbers of IL-6 expressing cells in the
ganglia, [3,9-11]. Expression of IFN- and TNF- transcripts persists
in TG latently infected with HSV strains unable to replicate in
neurons, indicating that neither HSV replication nor ability to
reactivate are required for persistent cytokine gene expression [3].
While CD4+ T cells appear to be important in immunized mice for
protection against challenge virus infection [12], CD8+ T cells appear
to be important for establishment of latent infection in mice [7]; and
CD8+ T cells specific for HSV persist in TG for long periods of time
[8]. Thus, there is evidence for long-term immune surveillance in the
ganglion during latent infection by HSV.

Chemokines are critical for recruiting inflammatory cells to infected
tissues. Chemokine specificity is due in large part to the
cell-specific expression of their respective receptors (reviewed in
[13-15]. Inflammatory-type receptors including CCR1, CCR2, CCR5, and
CXCR3 are expressed by activated T cells, macrophages, natural killer
(NK) cells, and immature (i.e. potent for antigen capture but not
antigen presentation) dendritic cells (DC), while homostatic-type
receptors including CCR7 and CXCR4 are highly expressed by resting T
and B cells and mature (i.e., antigen-presenting) DC (Table 1). In
addition, receptors including CCR2, CCR5 and CXCR3 are expressed on
cells (e.g. Th1 cells) specific for infection-induced inflammation,
while others including CCR3 and CXCR4 are on cells (e.g., Th2 T cells)
associated with allergic inflammation. Certain receptors are expressed
by specific subsets of a given cell type. For example, CCR6 is highly
expressed on Langerhans-like (CD34+) DC that migrate to skin, but not
on monocyte-derived DC that migrate to non-skin tissues (reviewed in
[14]. Acute viral infection in the mouse corneal model system is known
to induce the expression of cytokines and chemokines in corneal tissue.
Thomas et al. [16] observed the induction of transcripts encoding
N51/KC, macrophage inflammatory protein-1  (MIP-1), MIP-2 and
monocyte chemotactic protein 1 (MCP-1) and the cytokines IL-1, IL-6,
IL-12, and TNF-. Similarly, Tumpey et al. [17] showed induction of
MIP-2, MIP-1, and MCP-1 chemokines in the cornea during acute
infection. Infection of mouse fibroblast cells by HSV induces
expression of IL-6 [18], and infection of macrophages by HSV induces
RANTES expression directly [19]. Infection of other cell types may
induce expression of other cytokines and chemokines. Less is known
about chemokine expression during HSV latent infection phase. Halford
et al. [10] observed RANTES RNA expression, in addition to RNAs for
IL-2, TNF-, IFN-, and IL-10, during latent infection.

Recent studies have shown that HSV infection activates Toll-like
signaling and chemokine synthesis [20,21]. Thus, we hypothesized that
HSV infection might induce prolonged expression of a broad range of
chemokines at sites of acute and latent infection. Real-time
quantitative RT-PCR methods have facilitated studies of immune cell RNA
expression in mouse models [22,23]. We report here the use of real-time
RT-PCR to monitor RNA expression of selected chemokine receptors and
their chemokine ligands during HSV infection of mouse corneal and TG
tissue. Our data show that RNA encoding inflammatory-type chemokine
receptors and their ligands persists in infected corneas and TG long
after infectious virus can be detected, suggesting prolonged chemokine
production and subsequent homing of inflammatory immune cells to these
tissues. Strikingly, the data demonstrate the persistent expression of
chemokines and chemokine receptor genes in the apparent absence of
detectable viral productive infection transcripts in infected corneas.

Outline          Results

Abstract
Introduction
Results
Discussion
Materials and Methods
Competing interests
Authors' Contributions
Acknowledgments
References

Figures

Figure 1
HSV tk and ICP0 RNA expression in mock and HSV-infected cornea and TG

Figure 2
Relative levels of chemokine and chemokine receptor RNA expression in
mock and HSV-infected cornea

Figure 3
Relative levels of chemokine and chemokine receptor RNA expression in
mock and HSV-infected TG

Tables

Table 2
Primer and Probe Sequences

Table 3
Induction Ratio (HSV+/Mock) of Transcripts for Chemokine Receptors,
Chemokines and Cytokines in Cornea and Trigeminal Ganglia (TG)

Table 4
Induction Ratio (HSV+/Mock) of Transcripts for Chemokine Receptors and
Chemokines in Trigeminal Ganglia (TG) at Late Times Post-Infection

Development of TaqMan RT-PCR assays to measure viral and host gene
expression during acute and latent infection

To monitor RNA expression of viral and host genes during HSV infection
of mice, we developed TaqMan RT-PCR assays for the quantification of
transcripts from the HSV tk and ICP0 genes and from mouse genes
encoding selected chemokine receptors and their ligands. In the
real-time PCR assay detailed in Materials and Methods, RNA isolated
from corneal and ganglionic tissue was used for synthesis of cDNA.
Primers and Taqman probes for the viral or cellular genes (Table 2)
were used in real-time PCR assays to measure the concentration of cDNA
for each transcript.

To characterize the range over which the HSV tk and ICP0 real-time PCR
assays were accurate and linear, we tested 10-fold dilutions of
purified HSV genomic DNA (kind gift of Jean Pesola) starting from 5.5
=97 104 copies for tk and ICP0 gene levels. The HSV tk and ICP0
primer/probe sets gave linear amplification curves over 4 logs of
template concentrations until the limit of detection within the linear
range was reached at 55 DNA copies for tk and 550 copies for ICP0 (not
shown). At these limits of detection, the threshold cycle (CT) value,
which indicated the PCR cycle at which a significant increase in
amplification was first detected, was 39.2 for tk at 55 DNA copies and
36.5 for ICP0 at 550 DNA copies.

Using 2-fold dilutions of uninfected mouse TG cDNA, we observed that
the primer/probe sets for host genes listed in Table 2 including GAPDH
gave linear amplification curves over at least 3 and up to 7 dilutions.
In all cases, CT values changed by about 1 cycle for every 2-fold
change in template concentration as expected (not shown). Thus our
assays matched well with previously described TaqMan assays [22-24]
for linearity and sensitivity.

Following corneal inoculation of mice with HSV or virus diluent (mock),
we collected corneas and TG during acute (3 and 10 dpi) and latent (30
dpi) phases. To monitor viral gene expression in infected mice, we
tested tissue samples for tk and ICP0 gene transcripts. In infected
corneal tissue, HSV tk and ICP0 transcripts were readily detected at 3,
but not at 10 or 30 dpi where CT values = 40 (indicating no measurable
RNA) (Fig. 1). Thus we could not detect lytic transcripts in infected
corneas beyond the acute phase using this assay.

In infected TG, tk RNA peaked at 3 dpi then dropped precipitously
(200-fold) to low but readily detectable levels by 10 dpi. At 30 dpi,
we detected very low or undetectable tk RNA expression in infected TG.
In the experiment shown in Fig. 1A, we measured a CT value of 38.2 for
tk expression in infected TG at 30 dpi, resulting in a relative
expression value of 0.0002. In an independent experiment, we measured a
CT of 38.1 for tk RNA in 30 dpi TG; however, a CT value of 40 was
measured in two additional experiments (not shown). CT values for all
reactions without RT were 40, indicating no DNA contamination. Thus,
while tk expression in latent TG was at the limit of detection for our
assay, our ability to detect tk expression in some but not all latent
TG was consistent with previous reports in which very sensitive RT-PCR
assays were used to detect tk (and ICP0) gene transcripts in some but
not all TG during latent infection [5,25]. In those previous reports,
an assay that included a radioactive Southern blotting step subsequent
to RT-PCR could detect single copies of tk nucleic acid per PCR
reaction. Our present assay for tk transcripts is at least 50-fold less
sensitive than that used by Kramer and Coen [5].

ICP0 RNA levels were similar to tk in that they peaked at 3 dpi in
cornea and TG (Fig. 1B). However, because our ICP0 probe/primer set
overlaps latency-associated transcript minor (LAT)  coding
sequences, the signal detected at 10 and 30 dpi in TG but not cornea
may be due to minor LAT read-through RNAs. RT-PCR analysis of LAT
transcripts from the TGs at 30 dpi was consistent with latent virus in
infected TG (unpublished results).

Chemokine and chemokine receptor expression in infected cornea and
ganglia

We next used TaqMan RT-PCR to monitor expression of a selected series
of mostly T cell and macrophage-specific chemokine receptors and
chemokines in mock and HSV-infected cornea and TG. We chose chemokine
receptors CCR1, CCR2, CCR5, and CXCR3, which are expressed by activated
T cells, macrophages, NK cells, and immature DC that would be part of
the immune infiltration in response to HSV infection, and their ligands
MIP-1, MIP-1, RANTES, and MCP-1. For comparison, we included CCR3
which is primarily expressed on granulocytes, the CCR3 ligand
eotaxin-1, CCR6 which is primarily expressed on resting T cells and
immature Langerhans-like (i.e., skin homing) DCs, CCR7 which is
primarily expressed on resting T and B cells and mature DCs that home
back to lymphoid tissues, and CXCR4 which is broadly expressed on many
immune and non-immune cell types (Table 1). We also tested the
chemokine-inducing cytokines IFN- and TNF-, whose RNA and protein
have previously been shown to be expressed during both acute and latent
phases of HSV infection [3,9-11].

i. Chemokine and chemokine receptor expression in infected cornea

Epithelial cells of the cornea are the initial sites of replication
following infection but infectious virus and viral mRNAs are not
detectable past 710 dpi [26]. We harvested RNA from mock and
HSV-infected cornea at 3, 10, and 30 dpi, and tested for chemokine
receptor and chemokine RNA expression in parallel. As expected for
tissues supporting active replication or having recently cleared virus,
chemokine receptors CCR1, CCR2, CCR5, CCR7, CXCR3 and CXCR4, but not
CCR3 or CCR6, were highly expressed and strongly induced (i.e.,

MIP-1, RANTES, and MCP-1, but not eotaxin-1, were also highly
expressed and strongly induced in infected cornea at 3 and 10 dpi.
IFN- and TNF- were also induced in infected cornea as previously
reported [16]. Surprisingly, induction of all host RNAs tested
persisted into latent phase at 30 dpi in infected corneas. For example,
CCR1, CCR2, and CCR5 exhibited similar induction and similar or only
slightly reduced expression levels at 30 dpi as compared to earlier
time points. Relative expression and induction of CCR7 and CXCR4 in
infected cornea appeared to be biphasic in that values were high at 3,
lower at 10, and higher again at 30 dpi. These results suggested that
continued presentation of HSV antigens stimulates chemokine production
and subsequent homing of effector cells to cornea despite the apparent
clearance of infectious virus.

ii. Chemokine and chemokine receptor expression in infected ganglia

In infected TG, transcripts from the genes encoding receptors CCR1,
CCR2, CCR5, CCR7, and CXCR3 were induced by HSV infection during both
acute (3 and 10 dpi) and latent (30 dpi) phases (Fig. 3 and Table 3).
Peak induction of these RNAs was at 10 dpi during the clearance phase.
CXCR4 was induced at 10 and 30 dpi but not at 3 dpi. While we measured
induction of CCR3 and CCR6 at 10 and 30 dpi, their very low expression
was at the limit of our detection (i.e., relative expression values <
0.5) as also seen in corneas. RNAs for the MIP-1, MIP-1, RANTES,
and MCP-1 chemokines were also strongly induced at each timepoint,
particularly at 3 dpi. Eotaxin-1 was induced at 3 dpi, but much less so
at 10 and 30 dpi. As seen previously [3] cytokines IFN- and TNF-
were strongly induced at 3 and 10 dpi, but much less so at 30 dpi.

A striking finding in this analysis was the persistent expression of
inflammatory cell RNAs during the latent phase of TG infection when
detectable production of infectious virus has ceased. To determine if
induction of these RNAs persisted past 30 dpi, we monitored expression
of a limited number of transcipts from in TG collected at 45, 62, and
90 dpi. In previous studies [3-5], HSV genomic DNA was maintained at
constant levels (~104 copies per TG) for up to 150 dpi in infected TG,
indicating that latent virus persists well beyond 90 dpi in this mouse
model. Induction of all RNAs in our panel persisted for at least 62
dpi; furthermore, all but CCR3 and eotaxin-1 were also induced at 90
dpi (Table 4). Thus chemokine receptor and ligand expression persisted
long into the latent phase in infected TG.

Outline          Discussion

Abstract
Introduction
Results
Discussion
Materials and Methods
Competing interests
Authors' Contributions
Acknowledgments
References

Recent studies have shown that HSV infection induces Toll-like
signaling and chemokine synthesis. Thus, we hypothesized that HSV
infection might induce a broad range of chemokines at sites of primary
and latent infection. In agreement with and extending previous studies
[3,9-11], we have found evidence for persistent expression of
chemokines and trafficking of inflammatory cells including activated T
cells to acutely infected corneal tissue and to latently infected
trigeminal ganglia. We also observed prolonged expression of chemokine
and chemokine receptor gene transcripts in corneal tissue, the primary
site of HSV-1 infection in this model system, long after infectious
virus has been cleared. Microarray analysis of host gene expression has
also demonstrated long-term alterations of host gene expression during
latent infection by HSV, including alterations in expression of CXCR6
mRNA in TG [27]. These results argue for long-term persistence or
expression of viral antigens or immunogens and stimulation of
expression of these chemokines, even at the primary site of infection,
the cornea. Recent results [28] have shown similar elevated chemokine
expression in lung tissue after clearance of murine gamma herpesvirus
68. It will be of interest to determine how widespread this effect is
among different virus infections or whether it is unique to viruses
that persist in the host, such as the herpesviruses.

Potential mechanisms for elevated expression of chemokines and
chemokine receptors after viral clearance

Low level expression of viral lytic transcripts in TG during latent
infection has been documented [5], which could result in low level
expression of viral proteins. Recent results have shown that HSV-1 can
activate Toll-like receptor 2 to stimulate chemokine expression and
secretion and to activate NF-B regulated promoters [20]. Lund et al.
[21] showed that infectious HSV-2 and also purified HSV-2 DNA activates
signaling through DC-expressed Toll-like receptor 9, resulting in the
induction of IFN- secretion. Toll-like receptor activation by HSV-2
DNA raises the intriguing possibility that HSV DNA alone is at least
partially responsible for TLR-dependent induction of chemokine
expression in latent TG. Among the transcripts that we studied, we
detected persistent expression of transcripts for MIP-1, MIP-1, and
RANTES, whose expression is activated by Toll-like receptors [29].
Expression of MIP-1 and MIP-1 could recruit NK cells, which express
CCR5, and immature dendritic cells, which express CCR1 and CCR5, into
the site of infection. Thus, elevated expression of at least some of
the chemokines could be due to Toll-like receptor activation. It is
also possible that other chemokines that were not assayed in this or
previous studies are also induced during latent HSV infection via
Toll-like receptor dependent mechanisms. Elevated expression of
chemokine receptors is likely due to the chemokine-induced trafficking
of inflammatory cells to the site of infection or, in the case of 30
days postinfection or latent infection, the site of viral antigen
persistence.

Although we have not examined expression of IP-10, a chemokine also
induced by Toll-like receptor signaling [29], we did examine the
expression of transcripts for CXCR3, its receptor on activated T cells.
Levels of both are elevated during latent infection in TG. Thus,
stimulation of expression of this chemokine could attract activated T
cells to the latently infected TG, providing a mechanism for the
persistent presence of HSV-specific CD8+ T cells in latently infected
TG [8].

Implications of persistent chemokine expression

Long-term inflammatory responses in neural tissue could induce
pathology due to damage to neuronal cells. A number of neurological
diseases have been associated with HSV infection [30], and these could
be associated with these long-term inflammatory responses. In addition,
the possibility of other types of specific pathological effects is
raised.

Role of HSV in coronary heart disease

Recent data have shown an association between HSV-1 seropositivity and
myocardial infarction and coronary heart disease in older adults [31].
These authors hypothesized that HSV-1 reactivation from autonomic
nerves that innervate the coronary arteries could cause infection of
endothelial cells, endothelial injury, and the initiation of an acute
thrombotic event. Similarly, based on our work, HSV infection might
induce expression of MCP-1 and IL-8, which are known to cause adhesion
of monocytes to vascular endothelium [32], an early step in the
development of atherosclerotic lesions in mouse models (reviewed in
Gerszten et al. [32]. Therefore, the induction and prolonged expression
of these chemokines by HSV infection could play a role in the
pathogenesis of coronary heart disease.

Role of HSV in HIV transmission

Considerable evidence has accumulated for the role of genital herpes
infections in promoting the transmission of human immunodeficiency
virus (reviewed in [33]. Although we examined HSV-1 in these studies,
HSV-2 shares many biological properties with HSV-1. Thus, it is
conceivable that genital herpes infections could similarly induce the
expression of chemokines in the genital mucosae and the trafficking of
dendritic cells and CD4+ T cells to that site. In addition to the break
in the genital epithelium provided by the genital lesion, the
recruitment of dendritic cells and CD4+ T cells to sites of HSV
infection would provide cells to transport HIV to lymph nodes and the
primary host cell, respectively, and increase the potential for HIV
infection.

Implications for HSV biology and vaccine design

Recent studies on the persistence of CD8+ T cells in latently infected
ganglia have concluded that these cells play a role in maintaining the
latent infection [8]. The results presented here raise the possibility
that the presence of CD8+ T cells in latently infected TG's could be
the result of chemokine expression. Thus, further studies are needed to
establish the causal relationship between the presence of CD8+ T cells
in latently infected ganglia and maintenance of latent infection.

Various HSV strains, including replication-defective mutants and
amplicon vectors which do not establish neuronal latency efficiently,
have been shown to induce durable immune responses [12,34,35]. These
results suggest that the basis for the durable immune responses may be
the persistence of antigen or continued antigen expression at sites of
primary infection. Further studies are needed to determine the source
of this antigen and the mechanism of the induction of chemokine
expression at primary and latent sites of HSV infection.

http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?CMD=search&DB=pubmed
http://www.journals.uchicago.edu/CID/journal/issues/v26n3/mr57_541/mr57_541.web.pdf
http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=pubmed&doptstract&
list_uids613350
http://darwin.bio.uci.edu/~faculty/wagner/hsv7f.html

How does LAT influence HSV-1 latency and reactivation?

Experimental studies using viral regulatory mutants and cell activation
have shown that the need for; aTIF and a0 can be abrogated to some
degree by the induction of the early processes of cell division and
metabolic stress in non-replicating cells.  Thus, latency can be viewed
as a dynamic balance where one or a few latently infected cells
sporadically enter the early stages of viral gene expression as a
response to stress, and this process usually aborts without cell death
returning the cell to latent infection. Alternatively any limited virus
produced is eliminated by innate and adaptive immunity.

In order for successful reactivation to occur, then, not only must a
latently infected cell allow limited productive viral replication, but
also the host must be at an immunological "low point" where virus
recrudescence can proceed.  This view correlates well with the fact
psychological and physiological stress, known to be immuno-suppressive,
are potent inducers of HSV-1 reactivation.

http://jvi.asm.org/cgi/content/full/76/16/8003
http://microbiology.med.nyu.edu/mohr/mohr_res.shtml

The Interactions Between Herpes Viruses and Their Host Cells

My lab is interested in the interactions between herpesviruses and
their host cells. Over the past several years, we have concentrated on
those interactions important for the regulation of protein synthesis.

The lytic replication of many viruses requires that large quantities of
viral proteins be produced to assemble the next generation of viral
progeny. To accomplish this, viral functions must, i) prevent a
cellular antiviral response designed to globally inhibit cellular and
viral protein synthesis; and ii) ensure that viral mRNAs are translated
efficiently.

Figure 1. Zoom in
In the course of many different viral infections, copious amounts of
double stranded RNA are produced, activating the cellular kinase PKR.
Unchecked, activated PKR would phosphorylate eIF2 on its alpha subunit,
inactivating this critical translation initiation factor and hindering
virus assembly and replication by inhibiting protein synthesis. To
counteract this cellular response and prevent the cessation of protein
synthesis prior to the completion of the viral lifecycle, many viruses
encode factors that prevent the accumulation of phosphorylated eIF2
alpha. We have taken a molecular genetic and biochemical approach to
investigate herpesvirus encoded proteins that inhibit this antiviral
response. One of these proteins, the herpes simplex virus V1 gamma
34.5 gene product, binds a cellular phosphatase and dephosphorylates
the inactivated pools of phosphorylated eIF2. Another protein encoded
by the Us11 gene binds RNA via a novel RNA binding motif rich in
arginine and prolines (fig. 1). This RNA binding motif has a high
affinity for double stranded or structured RNA molecules. Currently, we
are exploring the molecular mechanisms by which these viral proteins
exert their effects. Significantly, both the Us11 and gamma 34.5 gene
products are important determinants of interferon resistance in primary
human cells infected with HSV-1.

Figure 2. Zoom in
To ensure that its mRNAs are translated, viruses must effectively
capture key, potentially limiting components of the host translational
machinery. One of these components is the multi-subunit eIF4F complex.
Composed of the cap binding protein eIF4E, the large molecular scaffold
eIF4G, and the eIF4A RNA unwinding enzyme, eIF4F recognizes the
7-methyl GTP cap structure at the 5 end of the mRNA and recruits
the 40S ribosome via an association between eIF3 and eIF4G (fig 2).
Finally, the cellular polyA binding protein (PABP), while itself not a
component of eIF4F, physically interacts with eIF4G through the PABP
interacting protein PAIP-1, bridging the 5 and 3 ends of
the mRNA and generating a circular topology which possibly serves as a
checkpoint to ensure the mRNA is both capped and polyadenylated prior
to translation initiation.

Figure 3. Zoom in
The activity of eIF4F is regulated in part by the 4E-BP1 translational
repressors that bind eIF4E and prevent it from associating with eIF4G.
Phosphorylation of 4E-BP1 by the cellular kinase mTOR results in the
release of free eIF4E (fig. 3), which can now in turn associate with
eIF4G and assemble the eIF4F complex. In addition, an eIF4F associated
kinase, mnk-1, binds eIF4G and phosphorylates eIF4E, stimulating
translation. Strikingly, phosphorylation of eIF4E together with 4E-BP1
is stimulated in resting cells infected with HSV-1, and 4E-BP1 appears
to be degraded by the cellular proteasome. Moreover, blocking eIF4E
phosphorylation with a specific mnk inhibitor dramatically reduces
viral replication. Finally, the assembly of eIF4F complexes is enhanced
considerably upon infection with HSV-1. All of these events appear to
be dependent upon the HSV-1 ICP0 gene product. Current work in the lab
is focused on understanding how this is achieved. It is likely that the
mobilization of eIF4F by viral functions is important for the virus to
replicate in quiescent, non-dividing cells, such as neurons.

A second area of interest concerns the design and use of attenuated
viruses as anti-tumor agents. Ideally, such a virus would be able to
replicate in and destroy cancer cells without destroying normal
terminally differentiated cells. Unfortunately, the attenuation
process, which is necessary to ensure safety, often substantially
impairs the ability of the virus to replicate in a variety of cells in
culture and in animals. We have modified an attenuated HSV-1 mutant by
genetic selection in cancer cells and isolated an attenuated variant
with enhanced ability to replicate in cultured tumor cells. Our
modified HSV-1 is an extremely potent anti - tumor agent in an animal
model of human cancer. Thus, additional mutations can be introduced
into the genome of weakened, attenuated viruses that confer upon them
enhanced anti Vtumor activity.

M2slo2...@nospam.invalid wrote:

inside

M2slo2...@nospam.invalid wrote:

inside