Herpes simplex virus infections can result in recurrent sores, stromal keratitis leading to blindness, or encephalitis. Current clinical strategies to address HSV infections target the productive life cycle of the virus. Nucleoside analogues, such as acyclovir and its derivatives, target viral DNA replication. However, nucleoside analogs have little effect on the latent state. Considerations of antiviral approaches targeting aspects of latency and reactivation must consider the mechanistic underpinnings for these phenomena. Latency is characterized by a severe reduction in transcription relative to productive infection. The repression seen during latency is due to the chromatinization the viral genome. The repressors that mediate and maintain chromatin on the viral genome are not known. These may serve as targets for the reactivation processes. It is also not known what viral and cellular mechanisms are involved in reversing the repressive effects of chromatin resulting in the resumption of productive gene transcription in the process of reactivation. We will address three hypotheses relevant to these issues: 1. Viral genomes are repressed by both constitutive and facultative heterochromatin and to be determined effectors of these chromatin types localize or bind to quiescent genomes to maintain these structures. 2. ICP0 is known to reverse the repressive effects of chromatin structure;however available evidence suggests that ICP4 and VP16 may also function to activate transcription in the face of epigenetic repression. We suggest that all three may be capable of further advancing the reactivation process. 3. ICP0 specifies multiple activities that contribute to the derepression of quiescent genomes. Three interconnected specific aims are proposed to address these hypotheses: 1. Determine the cellular components that are involved in the repression of quiescent and latent genomes. This will be addressed by a systematic ChIP and ChIP-seq analysis of quiescent and latent genomes, combined with shRNA depletion experiments. 2. Determine if and how the three viral transactivators, ICP4, ICP0 and VP16 function on genomes repressed by heterochromatin. We will provide ICP0, ICP4 and VP16 to cells harboring quiescent or latent viral genomes to assess the ability of these proteins to: i. reverse specific repressive chromatin structures, and ii. recrit RNA polII, and activate transcription. 3. Determine the binding partners of ICP0 involved in the reversal of repressive chromatin and the activation of transcription. Transdominant mutants of ICP0 will be TAP-tagged, and the purified ICP0-containing complexes will be analyzed by mass spectrometry to identify the cellular proteins contained in the complexes. The significance of these interactions will be genetically determined. It is anticipated that some of the identified proteins will be those present on repressed quiescent genomes identified in aim 1. The completion of these studies will provide a better understanding of the repression and activation of latent genomes.

Public Health Relevance

Antiviral drugs exist that target the productive cycle of HSV by acting as DNA replication inhibitors and do not target latency or the reactivation process. Reactivation of HSV-1 from latency is poorly understood. Understanding the mechanisms by which HSV persists in latent infection, and can reactivate, will facilitate interventions in these processes.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Special Emphasis Panel (ZRG1-IDM-M (02))
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Challberg, Mark D
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University of Pittsburgh
Schools of Medicine
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Dembowski, Jill A; DeLuca, Neal A (2018) Temporal Viral Genome-Protein Interactions Define Distinct Stages of Productive Herpesviral Infection. MBio 9:
Fox, Hannah L; Dembowski, Jill A; DeLuca, Neal A (2017) A Herpesviral Immediate Early Protein Promotes Transcription Elongation of Viral Transcripts. MBio 8:
Dembowski, Jill A; Dremel, Sarah E; DeLuca, Neal A (2017) Replication-Coupled Recruitment of Viral and Cellular Factors to Herpes Simplex Virus Type 1 Replication Forks for the Maintenance and Expression of Viral Genomes. PLoS Pathog 13:e1006166
Dembowski, Jill A; Deluca, Neal A (2017) Purification of Viral DNA for the Identification of Associated Viral and Cellular Proteins. J Vis Exp :
Harkness, Justine M; Kader, Muhamuda; DeLuca, Neal A (2014) Transcription of the herpes simplex virus 1 genome during productive and quiescent infection of neuronal and nonneuronal cells. J Virol 88:6847-61
Orzalli, Megan H; DeLuca, Neal A; Knipe, David M (2012) Nuclear IFI16 induction of IRF-3 signaling during herpesviral infection and degradation of IFI16 by the viral ICP0 protein. Proc Natl Acad Sci U S A 109:E3008-17
Ferenczy, Michael W; Ranayhossaini, Daniel J; Deluca, Neal A (2011) Activities of ICP0 involved in the reversal of silencing of quiescent herpes simplex virus 1. J Virol 85:4993-5002
Ferenczy, Michael W; DeLuca, Neal A (2011) Reversal of heterochromatic silencing of quiescent herpes simplex virus type 1 by ICP0. J Virol 85:3424-35
Ferenczy, Michael W; DeLuca, Neal A (2009) Epigenetic modulation of gene expression from quiescent herpes simplex virus genomes. J Virol 83:8514-24
Hadjipanayis, Costas G; Fellows-Mayle, Wendy; Deluca, Neal A (2008) Therapeutic efficacy of a herpes simplex virus with radiation or temozolomide for intracranial glioblastoma after convection-enhanced delivery. Mol Ther 16:1783-8

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