Inducible Cyclic AMP Early Repressor Produces Reactivation of Latent Herpes Simplex Virus Type 1 in Neurons In Vitro

Abstract

Herpes simplex virus type 1 (HSV-1) establishes a latent infection in neurons of the peripheral nervous system. During latent HSV-1 infection, viral gene expression is limited to latency-associated transcripts (LAT).HSV-1 remains latent until an unknown mechanism induces reactivation. The ability of the latent virus to periodically reactivate and be shed is essential to the transmission of disease. In vivo, the stimuli that induce reactivation of latent HSV-1 include stress, fever, and UV damage to the skin at the site of initial infection. In vitro, in primary neurons harboring latent HSV-1, nerve growth factor (NGF) deprivation or forskolin treatment induces reactivation. However, the mechanism involved in the induction of reactivation remains poorly understood. During latent herpes simplex virus type 1 (HSV-1) infection in sensory neurons, the viral genome is maintained in a nonreplicating state and viral gene expression is silenced, with the exception of the viral gene that encodes the latency-associated transcripts (LAT) (34). Reactivation of latent HSV-1 is induced by many different stimuli, including fever, stress, and UV irradiation or abrasion to the skin. Studies using LAT mutants indicate that LAT enhances the establishment of latency as well as the reactivation of latent 6,11,22,23,33). The signaling mechanisms controlling the induction of reactivation of latent HSV-1 are not yet understood.Cyclic AMP (cAMP) and nerve growth factor (NGF)-mediated pathways are involved in the induction of reactivation. Forskolin, chlorophenylthio-cAMP, or NGF deprivation results in reactivation of latent HSV-1 in primary neuronal cultures (29). Activation of these pathways is shown to result in phosphorylation and activation of the CRE-binding protein (CREB) (8, 9). Functional CREB response elements (CREs) have been identified within the LAT promoter at positions Ϫ85 and Ϫ43 from the site of transcription initiation (4,15,24). The CRE at Ϫ43 has been shown to be cAMP responsive in transient-transfection assays, and mutagenesis of this CRE results in reduced reactivation in rabbits latently infected with the recombinant virus (4). Characterization of the CRE at Ϫ85 is primarily limited to the observation that members of the CREB/ATF family can interact with the promoter in electrophoretic mobility shift assays (13, 17). Based on this evidence, it is possible that CREs in the LAT promoter may have a role in signaling that results in the reactivation of latent HSV-1.Previous studies have focused on activation of LAT transcription by signaling pathways (15,24). Based on the presence of elements in the promoter of LAT, the role of the inducible cAMP early repressors (ICER) in the induction of reactivation of latent HSV-1 was examined. The CRE modulator (CREM) gene family encodes transcriptional activators and repressors that are structurally related to the CREB/ATF family (26). The best-characterized CREM repressors are the ICER isoforms (18). ICER is a member of the basic-leucine zipper family and represses by virtue of i...

Histone deacetylase inhibitors induce reactivation of herpes simplex virus type 1 in a latency-associated transcript–independent manner in neuronal cells

“…Thus, LAT could be a viral response that attempts to protect the neuron during an event that induces apoptosis and/or reactivation (Gill and Windebank, 1998;Perng et al, 2000;Perng et al, 2001; Thompson and Sawtell, 2001). If LAT is serving a protective role during reactivation, a logical explanation would be the accumulation of LAT in cells that become protected and do not support reactivation (i.e., remain latently infected), whereas LAT would fail to accumulate to appreciable levels in cells supporting reactivation (Colgin et al, 2001;Halford et al, 1996). Because our analysis involved RT-PCR, it is not certain if the high levels of LAT are accumulating in the select few cells that reactivate virus, or lower levels are produced in the majority of cells that do not.…”

Tissue-Specific Splicing of the Herpes Simplex Virus Type 1 Latency-Associated Transcript (LAT) Intron in LAT Transgenic Mice

“…In sensory ganglia, LAT expression seems to be tightly regulated; during the lytic infection, most neurons express either the LAT or lytic genes, not both (21). In addition, there is a sharp drop in the amount of LAT during reactivation (3,30), with a decrease in transcriptional permissiveness of the LAT promoter occurring as early as 30 min postexplant (1). These observations, combined with genetic data describing LAT mutants as being defective in reactivation (2,10,13,16,24), have led to the model showing that the LAT RNA may play a role in transcriptional silencing of lytic genes during latency (2,7,15,36).…”

The 2-Kilobase Intron of the Herpes Simplex Virus Type 1 Latency-Associated Transcript Has a Half-Life of Approximately 24 Hours in SY5Y and COS-1 Cells

“…Nevertheless, a difference in the stability of the 2-kb LAT between cells in culture and acutely or latently infected cells in the peripheral nervous system cannot be ruled out. For instance, Colgin et al saw a rapid decrease in the number of neurons expressing LAT in cultured rat dorsal root ganglia induced to reactivate by either nerve growth factor (NGF) withdrawal or addition of forskolin (10), suggesting that LAT may have a shorter half-life in this context. However, those authors attribute this decrease to a downregulation of the LAT promoter and not to stability of the 2-kb LAT RNA itself.…”

“…A similar protein(s) may bind to the 2-kb LAT and influence debranching and/or the stability of the intron, and mutations such as Bam and Cons may affect the binding of this protein(s). Interestingly, if the half-life of the 2-kb LAT is actually similar in the cultured rat dorsal root ganglion reactivation model described above (10), then down-regulation of the LAT promoter alone would not account for the rapid decrease seen in LAT-positive neurons. Instead, the rapid decrease may indicate a role for an additional factor(s) to decrease the amount of LAT present.…”