Identification of DNA sequences in the latency related promoter of bovine herpes virus type 1 which are bound by neuronal specific factors

Bovine herpes virus 1 establishes a latent infection in sensory ganglionic neurons of cattle. During a latent infection, latency related (LR) transcripts are the only detectable viral RNAs. DNA sequences in the LR promoter are positively regulated by neural factors. The 5′ terminus of LR RNA in productively infected bovine cells is 20–30 nucleotides downstream from two overlapping TATA like elements. In contrast, the major start sites of LR transcription in trigeminal ganglia of latently infected cattle was 200–300 nucleotides upstream. Electrophoretic mobility shift assays (EMSA) were utilized to identify regions of the LR promoter that specifically bind factors present in dorsal root ganglia of cattle. Nuclear extracts from dorsal root ganglia of cattle or rat pheochromocytoma cells (PC12) contain abundant factors which specifically bind to a 72 bp XhoI-XbaI fragment. The 72 bp fragment is adjacent to the major start sites of LR transcriptional in trigeminal ganglia of latently infected cattle. In contrast, nuclear extracts from non-neural cells, bovine turbinate or Rat-2, did not exhibit similar binding patterns suggesting these factors were not abundant, had reduced binding affinity, or were absent in non-neural cells. Binding was localized to a 20 bp region of the XhoI-XbaI fragment by EMSA and Exonuclease III footprinting. When the XhoI-XbaI fragment was deleted, LR promoter activity was repressed in PC12 cells. Taken together, we conclude the XhoI-XbaI fragment is important for LR-RNA expression in neurons.