Analysis of sequences important for herpes simplex virus type 1 latency-associated transcript promoter activity during lytic infection of tissue culture cells

Morrow, John A.; Rixon, Frazer J.

doi:10.1099/0022-1317-75-2-309

Cited by 7 publications

(2 citation statements)

References 39 publications

(31 reference statements)

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“…That is, we hypothesize that the expression of LAT in neurons prevents progression to lytic infection when they are subsequently superinfected as the acute stage of infection proceeds. The regulation of the LAT gene is not completely delineated, but the promoter contains neuron-specific expression elements (2,3,9,46,89) and its strongest expression in terms of both intensity and number of neurons is during the peak days of acute virus replication in TG (44,59). These findings suggest that neurons that initially enter the latent pathway (e.g., that are infected with one or a few virions) begin to express the LAT-dependent establishment function.…”

Section: Discussionmentioning

confidence: 99%

Herpes Simplex Virus Type 1 Latency-Associated Transcript Gene Promotes Neuronal Survival

Thompson

Sawtell

2001

J Virol

215

264

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A complex interaction has evolved between the host's peripheral nervous system (PNS) and herpes simplex virus type 1 (HSV-1). Sensory neurons are permissive for viral replication, yet the virus can also enter a latent state in these cells. The interplay of viral and neuronal signals that regulate the switch between the viral lytic and latent states is not understood. The latency-associated transcript (LAT) regulates the establishment of the latent state and is required for >65% of the latent infections established by HSV-1 (R. L. Thompson and N. M. Sawtell, J. Virol. 71:5432-5440, 1997). To further investigate how LAT functions, a 1.9-kb deletion that includes the entire LAT promoter and 827 bp of the 5 end of the primary LAT mRNA was introduced into strain 17syn؉. The wild-type parent, three independently derived deletion mutants, and two independently derived genomically rescued variants of the mutants were analyzed in a mouse ocular model. The number of latent sites established in trigeminal ganglion (TG) neurons was determined using a single-cell quantitative PCR assay for the viral genome on purified TG neurons. It was found that the LAT null mutants established ϳ75% fewer latent infections than the number established by the parental strain or rescued variant. The reduced establishment phenotype of LAT null mutants was due at least in part to a dramatic increase in the loss of TG neurons in animals infected with the LAT mutants. Over half of the neurons in the TG were destroyed following infection with the LAT mutants, and this was significantly more than were lost following infection with wild type. This is the first demonstration that the HSV LAT locus prevents the destruction of sensory neurons. The death of these neurons did not appear to be the result of increased apoptosis as measured by a terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling assay. Animals latently infected with the LAT null mutants reactivated less frequently in vivo and this was consistent with the reduction in the number of neurons in which latency was established. Thus, one function of the LAT gene is to protect sensory neurons and enhance the establishment of latency in the PNS.A complex interaction between the virus and the host determines the fate of cells infected by herpes simplex virus type 1 (HSV-1). At the body surface, about 77 known viral lyticphase genes are actively transcribed during productive infection, which is invariably lethal for the cell. Latent infections are characterized by the down regulation of these lytic-phase genes in innervating sensory neurons of the trigeminal ganglia (TG), and these cells survive infection. The continued expression of the latency-associated transcript (LAT) is detectable by in situ hybridization in about 100 to 200 sensory neurons per mouse TG (for review, see references 1 and 87). The discovery of the LAT RNAs over a decade ago led to speculation that that they must play a central role in viral latency (70), but the function(s) of the LAT gene is not yet unde...

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Section: Discussionmentioning

confidence: 99%

Herpes Simplex Virus Type 1 Latency-Associated Transcript Gene Promotes Neuronal Survival

Thompson

Sawtell

2001

J Virol

215

264

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“…An important function of LATs is the inhibition of apoptosis, and although the mechanism is unclear, LATs do promote the production of Hsp70 that protects cells from stress (2). LATs are expressed in a varied and cell-type-specific manner in vivo and are not produced efficiently in quiescently infected rat neuronal or human fibroblast cultures (34,37,50). Therefore, in an attempt to mimic this state in cultures of human fibroblasts, we serum starved NHDFs (75,76) and preincubated these cultures at 41°C to generate nondividing human cell cultures expressing elevated levels of HSPs.…”

Section: Resultsmentioning

confidence: 99%

Efficient Quiescent Infection of Normal Human Diploid Fibroblasts with Wild-Type Herpes Simplex Virus Type 1

McMahon

Walsh

2008

J Virol

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Quiescent infection of cultured cells with herpes simplex virus type 1 (HSV-1) provides an important, amenable means of studying the molecular mechanics of a nonproductive state that mimics key aspects of in vivo latency. To date, establishing high-multiplicity nonproductive infection of human cells with wild-type HSV-1 has proven challenging. Here, we describe simple culture conditions that established a cell state in normal human diploid fibroblasts that supported efficient quiescent infection using wild-type virus and exhibited many important properties of the in vivo latent state. Despite the efficient production of immediate early (IE) proteins ICP4 and ICP22, the latter remained unprocessed, and viral late gene products were only transiently and inefficiently produced. This low level of virus activity in cultures was rapidly suppressed as the nonproductive state was established. Entry into quiescence was associated with inefficient production of the viral trans-activating protein ICP0, and the accumulation of enlarged nuclear PML structures normally dispersed during productive infection. Lytic replication was rapidly and efficiently restored by exogenous expression of HSV-1 ICP0. These findings are in agreement with previous models in which quiescence was established with HSV mutants disrupted in their expression of IE gene products that included ICP0 and, importantly, provide a means to study cellular mechanisms that repress wild-type viral functions to prevent productive replication. We discuss this model in relation to existing systems and its potential as a simple tool to study the molecular mechanisms of quiescent infection in human cells using wild-type HSV-1.Herpes simplex virus-1 (HSV-1) is a large, double-stranded DNA virus that establishes life-long latent infection in sensory neurons of affected individuals after spread at the primary site of infection (59). Lytic replication of the virus is well characterized and involves the temporally ordered expression of viral genes, broadly grouped into three classes, termed immediateearly, early, and late. The outcome of lytic replication is death of the host cell. However, during primary infection neurons that do not support lytic replication ultimately play host to the viral genome in a nonproductive state, termed latency, in which minimal transcription from the viral genome occurs. In response to certain environmental cues, such as UV light and stress, HSV-1 periodically reactivates from latency, replicating in ganglia and the differentiated epithelia that they innervate.The complex nature of latent infection means that it has proven challenging to study at the cellular level. Latency can be experimentally reproduced in animals, most commonly using rodents or rabbits, and these models have been used to develop a broad understanding of factors that regulate various aspects of the latent state (59). However, the inherent complexity of animal models means that they are largely unsuited to detailed mechanistic studies. To elucidate latency at the cellular ...

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Ebola and Marburg

Kobinger

Croyle²,

Feldmann

2009

Vaccines for Biodefense and Emerging and Neglected Diseases

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Analysis of sequences important for herpes simplex virus type 1 latency-associated transcript promoter activity during lytic infection of tissue culture cells

Cited by 7 publications

References 39 publications

Herpes Simplex Virus Type 1 Latency-Associated Transcript Gene Promotes Neuronal Survival

Herpes Simplex Virus Type 1 Latency-Associated Transcript Gene Promotes Neuronal Survival

Efficient Quiescent Infection of Normal Human Diploid Fibroblasts with Wild-Type Herpes Simplex Virus Type 1

Ebola and Marburg

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