Analysis of the latency-associated transcript/UL1-3.5 gene cluster promoter complex of pseudorabies virus

During latency, pseudorabies virus (PRV) DNA is preferentially retained in the neurons of the trigeminal ganglion and a spliced 8.5-kilobase poly-A RNA, designated large latency transcript (LLT), is synthesized. Because LLT is the only transcript made during the latent phase, the LLT promoter may be unique among all other PRV promoters that are active in productive infections. Organization of the PRV LLT promoter is quite complex because it coincides with the UL1-3.5 gene cluster promoter, but in the opposite orientation. By conventional designation, LLT is transcribed in the rightward direction while the UL1-3.5 gene cluster is transcribed in the leftward orientation. In this work, activities of the LLT promoter and the UL1-3.5 gene cluster promoter were investigated by transient reporter gene expression assay in cells of neuronal and non-neuronal origins. There are two TATA boxes in this region. We examined the promoter activities of the first TATA box with its 5' sequence (LAP1) and the second TATA box with its 5' sequence (LAP2). The UL1-3.5 promoter driven constructs gave no reporter gene activity in any of the experiments. Reporter gene activity was detected with LAP2 gene constructs, but not with LAP1 constructs, in both neuronal and non-neuronal cells. This is surprising because transcription of PRV LLT in vivo has been attributed to LAP1 and the initiation site was mapped downstream of the LAP1 TATA box and upstream of the LAP2 TATA box. Although LAP1 was not active in these experiments, there was a 3- to 10-fold enhancement of activity when LAP1 and LAP2 were placed in tandem.