Function of histone chaperones in HSV-1 chromatin sturcture during latency, establishing maintenance and reactivation

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 DESCRIPTION (provided by applicant): Herpes simplex virus (HSV) causes significant pain and suffering in humans. It is acquired by skin infection (typically mouth/lips for HSV-1 and genitals for HSV-2) where it replicates lytically and then spreads to the nervous system. It establishes a life-long latent infection within sensory nerve ganglia and persists as a double-stranded DNA (episome) within the nuclei of latently infected neurons. Periodically HSV reactivates from this latent state and causes cold sores (HSV-1) or genital lesions (HSV-2). The mechanism of how HSV establishes a latent infection or reactivates from latency is not understood. Over 50 million people in the US experience clinical recurrent HSV disease. HSV-1 is also the leading cause of infectious blindness in the US with over 30,000 new cases a year. While antiviral drugs can reduce the severity of symptoms, they do not block reactivation and they are not curative. Understanding how HSV interacts with the cell to regulate its lytic and latent life cycle could provide new targets for therapeutic intervention. Recent advantages in the field have demonstrated that transcriptional control of viral genes at the chromatin level plays a key role in the delicate balance between latency and reactivation, suggesting that chromatin is the key element in HSV latency. While the details of these regulatory processes are largely unknown, several recent findings have provided clues such as: evidence that HSV lytic genes are silenced by post-translational modifications of histones (epigenetic repression); insulator protein CTCF establishes epigenetic borders on viral DNA during latency, separating repressed lytic genes and active LAT region; nuclear protein DAXX and its interaction partner ATRX play a pivotal role by chaperoning the histone variants that can facilitate formation of mobile, transcriptionally active chromatin; DAXX and ATRX participate in intrinsic antiviral defenses repressing lytic infection. During latency establishing, DAXX complex participates in chromatin formation on HSV-1 DNA. In turn, this may create specific chromatin signature to establish proper loading and maintenance of CTCF at specific CTCF-binding elements of HSV genome thus creating epigenetic boundaries on this genome to regulate latency. These observations allowed us to formulate the main hypothesis of this proposal: the DAXX/ATRX-mediated loading of histone variants regulates dynamic deposition of CTCF on HSV-1 genome thus controlling appropriate chromatin formation that is critical for latency. We expect that the balanced deposition of histone variants represents a key necessary step in latency and predict that DAXX/ATRX complex plays an essential role in this process. In order to test this hypothesis we are combining the expertise of two co-investigators. Dr. Alexander Ishov is an expert in the DAXX/ATRX biology and in the analysis of intra-nuclear aspects of viral infection. Dr. David Bloom is an expert in epigenetic control of HSV gene expression and models to study HSV pathogenesis and latency. The key studies proposed in this project will investigate the involvement of DAXX/ATRX in recruiting histones variants to the regulatory elements of HSV genome and whether this alters the ability of the genome to establish and maintain latency via CTCF deposition in both in vitro and in vivo settings. These studies will determine whether DAXX/ATRX chaperone complex is involved in the regulatory balance between HSV lytic and latent infection and will provide essential data for the mechanistic understanding of this process. In addition to providing insight into the mechanism of HSV latency and reactivation, this highly innovative and focused pilot study has the potential to identify new therapeutic targets for treating HSV infections. If DAXX/ATRX were shown to have a role in regulating the switch between lytic and latent infection, this complex would be highlighted as target for anti-viral therapy.