Analysis of regulatory functions for the region located upstream from the latency-associated transcript (LAT) promoter of pseudorabies virus in cultured cells

The latency-associated transcript (LAT) promoter of pseudorabies virus (PrV) is unique among the many promoters of the viral genome in that it remains active during the latent state. The regulatory mechanism of PrV LAT gene expression is complex and different between latency and lytic infection of cultured cells. Although two different sequences, LAP1 and LAP2, are thought to be involved in LAT gene expression, the function of the upstream region of the LAT promoter (LAP1 and LAP2) remains an enigma, even in cultured cells. To analyze the function of the upstream region, it is necessary to examine the effects of the upstream sequence on LAT gene expression in the absence of other viral proteins. Transient expression assays were performed by employing a series of reporter plasmids in which various sequences upstream of the LAT promoter (from nucleotide positions -592 to +423 relative to the transcriptional start site of the large latency transcript (LLT)) were linked to the chloramphenicol acetyltransferase (CAT) gene in cells of neuronal and non-neuronal origin. We identified a region (from nucleotide positions -3606 to -1386) that was capable of repressing the LAT promoter activity in Vero cells by analyzing CAT gene expression of the series of reporter plasmids. This effect was not observed in Neuro-2a cells. We have also shown that the LAT promoter activity of the reporter plasmid containing the upstream region was repressed by the immediate-early gene product IE180 in Vero cells, but not in Neuro-2a cells. These results suggest that the upstream region of the LAT promoter may have a role in repressing LAT gene expression in cultured non-neuronal cells.