The long term goal of the proposed research is to define the molecular mechanisms involved in the transition from latent to lytic herpes simplex virus (HSV) gene transcription. They previously developed the hyperthermic stress (HS) reactivation model which is unique in that the production of infectious virus is detectable within 12-14 hours after the induction stimulus. Using this model, they have identified what is likely to be a key event in the regulation of reactivation, namely the up regulation of ICPO within 1 hour post HS. Insight into the molecular regulation of reactivation must ultimately be obtained from analysis of individual latently infected neurons. They have developed a new method, contextual expression analysis, CXA, to obtain quantitative information about the DNA and RNA in individual cells within solid tissues. In this proposal, the power of PCR and RT PCR will be harnessed through CXA to construct a molecular definition of latency and reactivation. Their ability to precisely quantify the number of latently infected neurons in the ganglia and examine the RNA and DNA content will allow them to meaningfully evaluate wild type and genetically engineered mutant strains to achieve the following specific aims: (1) Determine the impact of the number of latently infected neurons and/or the number of viral genome copies within individual latently infected neurons and/or the number of viral genome copies within individual latently infected neurons upon the initiation and progression of HS inducted reactivation in vivo; (2) Utilize CXA-RNA strategies to characterize viral transcription during latency and following HS induced reaction at the neuronal population and single cell level; (3) Determine the biological significance and biochemical basis of the rapid up regulation of the ICPO gene following HS induced reaction in vivo. Defining the regulatory mechanisms by which the """"""""latent"""""""" repository of viral genetic information periodically give rise to infectious virus is central to understanding this important aspect of the viral life cycle. Insight into these viral functions could contribute significantly toward our ability to design effective vaccines, develop treatments for the prevention of recurrent disease, and efficiently transfer, maintain and regulate foreign genes in the human host.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI032121-06A1
Application #
2470205
Study Section
Virology Study Section (VR)
Project Start
1992-01-01
Project End
2002-12-31
Budget Start
1998-01-01
Budget End
1998-12-31
Support Year
6
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Cincinnati Children's Hospital Medical Center
Department
Type
DUNS #
071284913
City
Cincinnati
State
OH
Country
United States
Zip Code
45229
Divanovic, Senad; Sawtell, Nancy M; Trompette, Aurelien et al. (2012) Opposing biological functions of tryptophan catabolizing enzymes during intracellular infection. J Infect Dis 205:152-61
Thompson, Richard L; Sawtell, Nancy M (2011) The herpes simplex virus type 1 latency associated transcript locus is required for the maintenance of reactivation competent latent infections. J Neurovirol 17:552-8
Sawtell, Nancy M; Triezenberg, Steven J; Thompson, Richard L (2011) VP16 serine 375 is a critical determinant of herpes simplex virus exit from latency in vivo. J Neurovirol 17:546-51
Thompson, Richard L; Sawtell, Nancy M (2010) Therapeutic implications of new insights into the critical role of VP16 in initiating the earliest stages of HSV reactivation from latency. Future Med Chem 2:1099-105
Thompson, Richard L; Preston, Chris M; Sawtell, Nancy M (2009) De novo synthesis of VP16 coordinates the exit from HSV latency in vivo. PLoS Pathog 5:e1000352
Holman, Holly A; MacLean, Alasdair R (2008) Neurovirulent factor ICP34.5 uniquely expressed in the herpes simplex virus type 1 Delta gamma 1 34.5 mutant 1716. J Neurovirol 14:28-40
Sawtell, N M; Thompson, R L; Haas, R L (2006) Herpes simplex virus DNA synthesis is not a decisive regulatory event in the initiation of lytic viral protein expression in neurons in vivo during primary infection or reactivation from latency. J Virol 80:38-50
Thompson, R L; Sawtell, N M (2006) Evidence that the herpes simplex virus type 1 ICP0 protein does not initiate reactivation from latency in vivo. J Virol 80:10919-30
Thompson, R L; Shieh, May T; Sawtell, N M (2003) Analysis of herpes simplex virus ICP0 promoter function in sensory neurons during acute infection, establishment of latency, and reactivation in vivo. J Virol 77:12319-30
Sawtell, N M (2003) Quantitative analysis of herpes simplex virus reactivation in vivo demonstrates that reactivation in the nervous system is not inhibited at early times postinoculation. J Virol 77:4127-38

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