Role of anisomorphic DNA conformations in the negative regulation of a herpes simplex virus type 1 promoter.

The a sequence is a bifunctional element in the herpes simplex virus type 1 (HSV-1) genome which possesses both the signals required for the cleavage and encapsidation of replicated viral DNA and the promoter-regulatory sequences for the gene encoding the viral neurovirulence factor ICP34.5. Since the ICP34.5 promoter lacks features that are characteristic of most HSV-1 promoters, including a canonical TATA box, an initiator element, and upstream binding sites for host cell transcription factors, a mutational analysis was undertaken to identify the cis-acting elements which mediate transcription of this gene in transient transfection assays. A deletion derivative containing sequences just 83 nucleotides upstream of the second of two cap sites was found to exhibit full promoter activity. However, the presence of either of two far upstream regions, which coincided with the DR2 and DR6 tandem GC-rich repeat arrays, acted to abrogate transcriptional activity both in this segment of the ICP34.5 promoter and in a heterologous promoter construct. The DR2 and DR6 repeat arrays each possessed an unwound S1 nuclease-sensitive DNA conformation (anisomorphic DNA) whose formation was shown to be critical for mediating this transcriptional repression effect. Moreover, results from in vivo titration experiments suggested the existence of a cellular protein(s) which can mediate transcriptional repression in the ICP34.5 promoter by specifically interacting with the single-stranded regions of these tandem repeat arrays. Such DNA conformation-dependent transcriptional silencing appears to represent a novel mechanism of gene regulation in the HSV-1 life cycle.