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Review
. 2022 Jun 2;14(6):1209.
doi: 10.3390/v14061209.

Impact of Cultured Neuron Models on α-Herpesvirus Latency Research

Angus C Wilson  1
Affiliations
Review

Impact of Cultured Neuron Models on α-Herpesvirus Latency Research

Angus C Wilson. Viruses. .

Abstract

A signature trait of neurotropic α-herpesviruses (α-HV) is their ability to establish stable non-productive infections of peripheral neurons termed latency. This specialized gene expression program is the foundation of an evolutionarily successful strategy to ensure lifelong persistence in the host. Various physiological stresses can induce reactivation in a subset of latently-infected neurons allowing a new cycle of viral productive cycle gene expression and synthesis of infectious virus. Recurring reactivation events ensure transmission of the virus to new hosts and contributes to pathogenesis. Efforts to define the molecular basis of α-HV latency and reactivation have been notoriously difficult because the neurons harboring latent virus in humans and in experimentally infected live-animal models, are rare and largely inaccessible to study. Increasingly, researchers are turning to cultured neuron infection models as simpler experimental platforms from which to explore latency and reactivation at the molecular level. In this review, I reflect on the strengths and weaknesses of existing neuronal models and briefly summarize the important mechanistic insights these models have provided. I also discuss areas where prioritization will help to ensure continued progress and integration.

Keywords: HSV-1; HSV-2; VZV; cell culture; latency; neurons; reactivation.

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Conflict of interest statement

The author declares no conflict of interest. The sponsors had no role in the conceptualization or writing of the manuscript.

Figures

Figure 1
Figure 1
Randy Cohrs in his element reading out discussion topic suggestions in front of a roaring fire on the last night of CALS 2017. Photograph generously provided with permission by CALS.
Figure 2
Figure 2
Host kinase AKT integrates NGF signaling with maintenance of chromosomal integrity to maintain HSV-1 latency. Studies in rat SCG neurons have shown that the AKT integrates external signaling from NGF interacting with its cognate receptor TrkA and intracellular DNA damage signaling initiating at topoisomerase 2 (Topo2)-induced DNA breaks. By shuttling between the cytoplasm and nucleus, AKT is kept in an active state through simultaneous phosphorylation of threonine-308 (T308P) by PDK1 in the cytoplasm and of serine 473 (S473P) by DNAPK in the nucleus. The AKT-mTORC1 axis ensures continuous cap-dependent protein synthesis, which is required to maintain the HSV-1 genome in a transcriptionally repressed state. Silencing of viral productive cycle genes involves the combined activities of repressive facultative heterochromatin chromatin, microRNAs, and potentially by lncRNAs such as the viral latency-associated transcript (LAT).
Figure 3
Figure 3
Biphasic model of HSV-1 reactivation. Studies using the rat and mouse SCG neuron infection models have found that HSV-1 reactivation proceeds through two mechanistically distinct steps referred to as Phase I and Phase II. Stresses such as interruption of growth factor signaling activates dual leucine zipper kinase (DLK) which in turn activate JNK resulting in phosphorylation of neuronal transcription factors and the repressive chromatin associated with the viral genome. JNK targets include serine-10 of histone H3 (H3S10P), which is adjacent to methylated lysine-9 (H3K9me3), a mark of repressive chromatin. This combinatorial mark (‘methyl/phospho switch’) renders chromatin permissive to RNAPII transcription, allowing widescale but low-level expression of viral productive cycle mRNAs. This transient animation of the viral transcriptome is termed Phase I. Although not yet test directly, dephosphorylation could potentially return animated genomes to their original transcriptionally silent state. The viral regulator VP16 is synthesized during Phase I but accumulates in the cytoplasm and does not influence viral gene expression. However, in a few neurons, VP16 is transported into the nucleus together with the coactivator HCF-1 and together recruit additional chromatin modifiers that remove repressive modifications and likely add activating marks such as methylation on lysine-4 of histone (H3K4me3) and/or acetylation of lysine 27 (H3K27ac) rendering the chromatin fully permissive for transcription. This VP16-dependent step is termed Phase II and results in higher levels of productive cycle gene transcription and permits the viral DNA genome amplification, which is not detected in Phase I.
Figure 4
Figure 4
Subcellular localization of the Gadd45b protein acts as a marker of HSV-1 reactivation heterogeneity within a neuronal population. Schematic showing the different patterns of Gadd45b protein (purple) accumulation within the cell bodies of latently-infected rat SCG neurons treated with reactivation inducer LY294002. Protein levels are elevated only in latently infected neurons and is distributed throughout the cytoplasm and nucleoplasm. In a small number of neurons, Gadd45b is excluded from the nucleus and these neurons contain EdU-positive foci (red) corresponding to sites of active viral DNA replication (vRC) a marker of Phase II. These individual neurons are considered to be engaged in successful reactivation. In another rare population, Gadd45b accumulates as discrete puncta with the nucleus. These neurons are always EdU-negative and likely have not entered into Phase II and represent unsuccessful or abortive reactivation events. For full details see Hu et al., 2021 [45].
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