English, C.; (2006) Molecular characterisation of the Herpes Simplex Virus-1 LATP2 enhancer region. Doctoral thesis , University of London.

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Abstract

HSV1 vectors have previously been produced with the ability to direct long-term expression of a transgene within the nervous system. These viruses have an arrangement of the latency promoter and enhancer regions LAP1 and LATP2 such that LATP2 exerts long-term expression onto LAP1 and an exogenous promoter at the same time, in a back-to-back fashion. To characterise the LATP2 region, two series of vectors were produced containing deletion mutations of the region placed in different orientations to LAP1 within the context of the original vectors. The vectors were tested in vitro and in vivo in the PNS and a potentially repressive region within LATP2 was identified. The enhancer activity of the region was also localised to a defined area. As the HSV1 genome is associated with histones and modification of these is a method of transcriptional control, histone modification could be one mechanism that the virus uses to keep the LAT region active during latency. This was investigated by examining the acetylation of histones associated with the LAT region, including LATP2, at lytic and latent timepoints in an in vitro system by ChIP assay. These studies found that although no significant differences in acetylation at different loci of LATP2 was found, the LAT regulatory region generally appears to be more associated with hyperacetylated histones during latency than non-LAT promoters and that this increased acetylation is conferred onto an exogenous promoter when placed within the LAT region. The findings in this thesis should provide insight into the functioning of the LAT region and may allow the development of improved HSV1 vectors for gene therapy in the nervous system.

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