The contrasting infection strategy of herpes simplex virus (HSV) consists of an initial primary lytic infection in epithelial cells, followed by establishment of lifelong latency in sensory and autonomic neurons of the peripheral nervous system that innervate the site of infection. Any cellular stress trigger, ranging from external stimuli such as UV radiation or nerve injury to psychological and physiological stress, can reactivate HSV from latency in the neurons, resulting in recurrent disease episodes. Stress hormones and deprivation of neurotrophic factor (NTF) both have a strong correlation with HSV reactivation from neurons. However, neuronal signaling pathways cardinal to HSV latency and reactivation are still not clear. This dissertation provides new understanding of HSV latency and reactivation in response to two orthogonal stress stimuli, viz. stress hormones epinephrine (EPI) and corticosterone (CORT), as well as NTF deprivation that simulates a nerve injury in primary neuronal cultures.
In this dissertation, we demonstrate that physiological stress hormones EPI and CORT differentially regulate HSV-1 and HSV-2 reactivation in adult neurons. Both EPI and CORT treatment reactivated only HSV-1 in sympathetic superior cervical ganglia (SCG) neurons, while HSV-2 was reactivated only by CORT in both sensory trigeminal ganglia (TG) neurons and sympathetic superior cervical (SCG) neurons. EPI utilized the combination of α and β adrenergic receptor complex, while CORT signaled through glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) to reactivate HSV in the neurons. NTFs are tissue-target derived growth factors required for neuronal protection and survival. Neurotrophins are also required for maintaining HSV latency, as NTF deprivation reactivates both HSV-1 and HSV-2 in adult sensory TG and sympathetic SCG neurons. In addition, assessing the temporal kinetics of HSV gene expression showed differential expression profiles of viral immediate-early (IE) genes ICP0, ICP4, ICP27 and trans-activator VP16 following treatment with stress hormones and NTF deprivation in HSV-1 and HSV-2 infected neurons.
We also show that different molecular mechanisms are involved in HSV latency and reactivation, which are dependent on the stimuli and the type of neurons. Tyrosine kinase receptor-mediated PI3K-Akt-mTORC signaling cascades have been studied for their role in maintaining HSV latency. Activation of β-catenin signalosome expression has also been implicated during HSV latency and following reactivation. GSK3β is a key effector molecule that inter-connects Akt and β-catenin mediated pathways, forming an Akt-GSK3β-β-catenin signaling axis. Analyzing the Akt-GSK3β-β-catenin signaling in response to stress hormone and NTF deprivation revealed significant differences in protein expression levels between HSV-1 and HSV-2 infected sensory and sympathetic neurons. In HSV-1 infected neurons, the Akt-GSK3β-β-catenin maintains the signal transmission in order to keep the neurons alive, but HSV-2 infections obliterated the entire axis in the adult neurons, particularly in sympathetic neurons. In summary, we demonstrate that HSV-1 and HSV-2 do not have a 'one for all' infection mechanism. Establishment of latency and reactivation by HSV is virus specific, stimulus specific and neuron specific. / Doctor of Philosophy / Herpes simplex viruses (HSVs) are common global viral pathogens that are responsible for causing lifelong painful infections and debilitating disease. The two serotypes of HSV include HSV-1, which is associated with oral or ocular disease but can also cause genital disease, and HSV-2, which is predominantly associated with genital herpes. Once infected, both HSV-1 and HSV-2 are present as lifelong reservoirs in our peripheral neurons.
Stress stimuli mediated by our stress hormones or external triggers, such as nerve trauma or an axonal injury, can periodically reactivate the latent virus to cause recurrent disease. Clinical manifestation of HSV recurrences range from asymptomatic viral shedding to painful blisters, cold sores, or herpetic keratitis. In some cases, the virus can spread to the central nervous system, causing encephalitis or recurrent meningitis. No vaccines have been approved yet, and the current treatment utilizes nucleoside analogs, such as acyclovir and its prodrug valacyclovir, to ameliorate the symptoms of HSV infection by halting viral replication and if taken as a daily prophylaxis, reduces the chances of clinical recurrence. Given the route and transmission efficiency of HSV, it is practically impossible to prevent herpes infection. To develop strategic therapeutic interventions to lock the virus in its latent phase in the neurons and prevent it from reactivation, a better understanding of neuronal signaling pathways cardinal to HSV latency and reactivation is necessary. However, neuronal signaling pathways cardinal to HSV latency and reactivation are still not clear.
In this dissertation, we make contributions to better understand HSV latency and reactivation in response to stress stimuli. We show that different stress stimuli exert preferential reactivation between HSV-1 and HSV-2, and are further dependent upon the neurons where they establish latency. Our study specifically focuses on three neuronal stressors that have been associated with HSV recurrences: two stress hormones, epinephrine (EPI) and corticosterone (CORT), as well as deprivation of neurotrophic factors (NTF) that simulates nerve injury. We also focused on a neuronal signaling cascade involved in the response to all of these stimuli, Akt-GSK3β-β-catenin, and viral gene transcripts that respond to these stimuli during reactivation. Comprehensive understanding of the neuronal processes and viral gene transcripts involved during HSV-1 and HSV-2 reactivation in neurons will help the herpes virology field towards development of targeted therapies and vaccines to prevent reactivation and recurrent disease.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/111559 |
| Date | 18 August 2022 |
| Creators | Goswami, Poorna |
| Contributors | Graduate School, Bertke, Andrea S., Caswell, Clayton Christopher, Hodes, Georgia E., Duggal, Nisha K. |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Format | ETD, application/pdf, application/pdf, application/pdf, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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