In Vivo Knockdown of the Herpes Simplex Virus 1 Latency-Associated Transcript Reduces Reactivation from Latency

Abstract

During herpes simplex virus (HSV) latency, most viral genes are silenced, with the exception of one region of the genome encoding the latency-associated transcript (LAT). This long noncoding RNA was originally described as having a role in enhancing HSV-1 reactivation. However, subsequent evidence showing that the LAT blocked apoptosis and promoted efficient establishment of latency suggested that its effects on reactivation were secondary to establishment. Here, we utilized an adeno-associated virus (AAV) vector to deliver a LAT-targeting hammerhead ribozyme to HSV-1-infected neurons of rabbits after the establishment of HSV-1 latency. The rabbits were then induced to reactivate latent HSV-1. Using this model, we show that decreasing LAT levels in neurons following the establishment of latency reduced the ability of the virus to reactivate. This demonstrates that the HSV-1 LAT RNA has a role in reactivation that is independent of its function in establishment of latency. In addition, these results suggest the potential of AAV vectors expressing LAT-targeting ribozymes as a potential therapy for recurrent HSV disease such as herpes stromal keratitis, a leading cause of infectious blindness. Herpes simplex virus (HSV) establishes a lifelong infection and remains dormant (latent) in our nerve cells. Occasionally HSV reactivates to cause disease, with HSV-1 typically causing cold sores whereas HSV-2 is the most common cause of genital herpes. The details of how HSV reactivates are largely unknown. Most of HSV's genes are silent during latency, with the exception of RNAs made from the latency-associated transcript (LAT) region. While viruses that make less LAT do not reactivate efficiently, these viruses also do not establish latency as efficiently. Here we deliver a ribozyme that can degrade the LAT to the nerve cells of latently infected rabbits using a gene therapy vector. We show that this treatment blocks reactivation in the majority of the rabbits. This work shows that the LAT RNA is important for reactivation and suggests the potential of this treatment as a therapy for treating HSV infections.

The etiology of Bell’s palsy: a review

“…After the establishment of HSV-1 latency, an adeno-associated virus delivered a LAT-targeting hammerhead ribozyme to rabbit neurons infected with HSV-1. Using this model, decreasing LAT levels in neurons reduced the ability of the virus to reactivate [39]. This suggests the potential for reverse validation of BP etiology with viral reactivation, as well as a potential avenue for the treatment of HSV infection.…”

“…In patients with higher 6-month HB score; their IAC inlet and mid-canal values were lower Table 2 Summary of key evidence for the etiological theory about virus infection Key references Summary of evidence [21][22][23][24] The α-HV which target peripheral neurons (e.g., HSV-1, HSV-2, and VZV) can establish lifelong infections and infectivity potential in the host including in the autonomic and sensory ganglia of the head, neck and cranial [25] Reactivation of HSV-1 centered around the geniculate ganglion was first outlined by McCormick in 1972 [26] The presence of HSV-1 deoxyribonucleic acid (DNA) was detected in clinical specimens, i.e., intra-temporal facial nerve endoneural fluid in Bell's palsy patients [27][28][29] Animal models have the capability to cause facial paralysis through initial infection and virus reactivation incited by immune modulation [36,37] Earlier work examining cellular electrophysiology in the setting of herpes infection demonstrated a pathway for the quick and dynamic control of excitability in sensory neurons by internalization of sodium channels. The processes of intra-axonal degeneration would drive the abrupt onset of Bell's palsy [38] The aquaporin 1 water channel protein (AQP1) in Schwann cells of intratemporal facial nerve is involved in the evolution of facial palsy caused by HSV-1 and may play an important role in the pathogenesis of this disease [39] Decreasing LAT levels in neurons reduced the ability of the virus to reactivate. This suggests the potential of reverse validation of bell's palsy as a virus reactivation Table 3 Summary of key evidence for the etiological theory about ischemia Key references Summary of evidence [46] Endoneurial blood supply to peripheral nerves is not uneven.…”

Blockade of PD-1 and LAG-3 Immune Checkpoints Combined with Vaccination Restores the Function of Antiviral Tissue-Resident CD8+TRMCells and Reduces Ocular Herpes Simplex Infection and Disease in HLA Transgenic Rabbits

“…At the end of the acute phase, HSV-1 travels up sensory neurons to the trigeminal ganglia (TG), where it establishes lifelong latency in its host (7)(8)(9)(10)(11). Reactivation of latent virus from neurons of the TG, anterograde transportation to nerve termini, and reinfection of the cornea can cause potentially blinding keratitis and is the major issue with HSV-1 infection globally (12)(13)(14)(15).…”

CD28 Costimulation Is Required for Development of Herpetic Stromal Keratitis but Does Not Prevent Establishment of Latency

“…Previous studies have shown that there is a significant influx of CD8 ϩ T cells in the TG of infected mice (26)(27)(28). When we analyzed CD8 ϩ T cells isolated from infected TG, we found that CD28KO mice initially displayed significantly fewer CD8 ϩ T cells at days 7 and 14 but that by day 41 there were no significant differences between CD28KO and wild-type mice.…”

“…*, P values ranged from 0.01 to 0.005. hybridization (FISH) analysis to quantitate the number of neurons expressing LAT as a means of quantifying latent infection with HSV-1 in infected TG. Previous studies examining HSV-1 latency have shown that using a similar methodology detects approximately 25% to 30% of latently infected ganglionic neurons (26)(27)(28). We anticipated that the results of that analysis would parallel the genome copy numbers, namely, that B6-CD28KO mice would display significantly greater numbers of latently infected cells.…”