Identification of a major regulatory sequence in the latency associated transcript (LAT) promoter of herpes simplex...

The latency associated transcript (LAT) gene is the only viral genomic region that is abundantly transcribed during herpes simplex virus type 1 (HSV-1) neuronal latency. As such, it may play an important role in HSV-1 latency and/or reactivation. The regulation of the LAT gene is complex and appears to include a combination of positive and negative functional elements in and near the LAT promoter. In this study, transient CAT assays were used to map the minimal promoter necessary for constitutive activity in neuronal and nonneuronal cells to between nucleotide positions -161 and -2 (relative to the start of LAT transcription). The region from -283 to -161 was able to slightly increase promoter activity of the minimal promoter and appeared to have a larger effect in neuronal derived cells. Gel-shift experiments using nuclear extracts from neuronal and nonneuronal derived cells detected a major factor that bound specifically to the -161 to -2 probe. We designated this factor LAT promoter binding factor (LPBF). Two additional minor factors also bound specifically to the minimal promoter. DNase I footprint analysis and gel-shift competition experiments demonstrated that LPBF bound to a region that includes the palindromic sequence CCACGTGG located at nucleotides -72 to -65. Deletion of this palindrome resulted in a loss of binding of LPBF from the minimal promoter region and an 8- to 30-fold reduction in promoter activity in both neuronal and nonneuronal cells. Thus, LPBF appears to play a major role in LAT promoter regulation.