Most of the human population world-wide has been infected by herpes simplex viruses. Following the initial lytic infection, HSVs establish permanent latent infections within sensory neurons. Reactivation of latent virus not only results in viral disease (new infections, blindness, and encephalitis) but also contributes to HIV infection, diabetes, cardiovascular and neurodegenerative diseases. No effective vaccine is available and no therapy eliminates latency or prevents reactivation. The long-term goal of this project is to find interventions for recurrent HSV episodes by defining mechanisms that control establishment and reactivation of HSV-1 latency. The gene expression cascade during HSV-1 lytic infection begins with activation of immediate-early (IE) gene transcription by the virion protein VP16 with host factors Oct-1 and HCF-1. In contrast, the initial events in the reactivation from latency are still poorly defined. The central hypothesis of this proposal is that regulation o both VP16 expression and activity underlie the establishment of latency and reactivation from latency. These two levels of control involve multiple positive and negative inputs to allow or inhibit viral replication in the sensory neuron in vivo.
Aim 1. This project will determine the mechanism of de novo VP16 gene activation and silencing in sensory neurons in vivo. The working hypothesis is that the VP16 gene in the HSV-1 genome can be regulated by action of neuron-specific and stress-responsive promoter elements and corresponding transcription factors either to allow or inhibit lytic replication upon initial infection or exit from latency. Uing recombinant viruses in a mouse model of infection and latency, we found that expression of VP16 is both necessary and sufficient to trigger the exit from latency and we identified a neuron-specific promoter for the VP16 gene. We will test whether this promoter controls the entry into lytic phase infection during acute infection and during reactivation. We will determine whether the predicted transcription factors bind to the various elements of this promoter to positively or negatively regulate VP16 gene expression, whether singly, in combination, or in competition.
Aim 2. This project will define the VP16 coactivator interactions essential for VP16-dependent exit from latency and identify mechanisms regulating these interactions in vivo. Our data strongly suggest that the exit from latency by HSV is regulated by CK2 mediated phosphorylation and that this phosphorylation may be also competitively regulated by O-GlcNAcylation (a PTM that regulates signaling in response to nutrients and stress). Our goal is to elucidate the functions of each of these PTMs in viral latency and to define the roles of their crosstalk in regulating immediate early gene expression of viral proteins through VP16 transactivation. The outcomes of this work will identify transcription factors or protein modifying enzymes that could be targets for future development of therapeutic interventions for HSV reactivation.
Herpes simplex virus causes serious disease in humans worldwide. Viral latency and reactivation in the nervous system are central to the disease process and VP16 is a key regulator of these events. We have identified a novel interaction between a major host cell nutrient/stress sensor and VP16 function. The proposed research will identify how this interaction regulates viral reactivation from latency. This knowledge will lead t novel antiviral targets and the improved design of treatment and prevention strategies.
|Thompson, Richard L; Williams, Robert W; Kotb, Malak et al. (2014) A forward phenotypically driven unbiased genetic analysis of host genes that moderate herpes simplex virus virulence and stromal keratitis in mice. PLoS One 9:e92342|