Epstein-Barr virus (EBV) infects 95% of the human population and contributes to 1-2% of all human cancers. To date, there are no specific therapies to eradicate cancers associated with this virus. The successful development of EBV-specific anti-cancer strategies requires an in-depth understanding of the molecular mechanism controlling viral latency and reactivation. Currently, there are fundamental gaps in our understanding of how cellular factors control EBV latency and reactivation. The central goal of this research program is to elucidate the role of protein inhibitor of activated STAT1 (PIAS1) in the EBV life cycle. Our recent study revealed that PIAS1 is an EBV restriction factor that inhibits virus reactivation from latency. We have demonstrated that PIAS1 is cleaved via caspases and phosphorylated by an EBV protein kinase upon lytic induction and that this is required for EBV replication. The mechanism for controlling lytic EBV replication is related to the ability of PIAS1 to inhibit the transcriptional activity of viral and cellular transcription factors critical for lytic gene expression. In addition, PIAS1 also regulates the expression of a group host genes involved in the cholesterol biosynthesis process, a pathway important for EBV lytic replication. These results lead to our central hypothesis that PIAS1 restricts EBV lytic replication by inhibiting viral and cellular transcription factors, and that PIAS1 is antagonized by caspase-dependent cleavage and viral kinase-mediated phosphorylation to foster lytic replication. Towards this hypothesis, we propose to pursue two Specific Aims:
AIM 1 : To determine the mechanisms by which PIAS1 suppresses EBV lytic replication. We will identify and validate PIAS1 binding sites on the EBV and host genomes in infected cells. We will determine how PIAS1 regulates EBV lytic gene expression via inhibiting viral and cellular transcription factors. We will also determine how PIAS1 inhibits EBV reactivation partially through modulating the cholesterol biosynthesis pathway.
Aim 2 : To elucidate the mechanisms by which PIAS1-mediated suppression is antagonized to facilitate EBV lytic replication. PIAS1 is cleaved by caspase-3, -6 and -8, and phosphorylated by an EBV protein kinase during lytic replication. We will identify PIAS1 phosphorylation sites using proteomic and molecular approaches. We will elucidate the interplay between the phosphorylation and cleavage PIAS1 that facilitates EBV replication. Our proposed research will significantly improve our understanding of how PIAS1, as a restriction factor, regulates EBV infection and how PIAS1 is regulated by viral and cellular factors during reactivation. These studies will have the potential to illuminate the molecular basis of EBV lytic reactivation controlled by one central host factor.

Public Health Relevance

Epstein-Barr virus (EBV) is etiologically linked to several human malignancies including nasopharyngeal carcinoma, lymphoma and a subtype of gastric cancer. This project investigates the key role of PIAS1 in regulating EBV latency and reactivation. The proposed study will yield new insight into virus-host interactions that modulate the EBV life cycle and facilitate the development of novel therapeutic strategies for eradicating EBV-associated disease.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Virology - A Study Section (VIRA)
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Natarajan, Ramya
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Virginia Commonwealth University
Schools of Dentistry/Oral Hygn
United States
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