2 HERPES-SIMPLEX VIRUS TYPE-1 LATENCY-ACTIVE PROMOTERS DIFFER IN THEIR CONTRIBUTIONS TO LATENCY-ASSOCIATED TRANSCRIPT EXPRESSION DURING LYTIC AND LATENT INFECTIONS

Herpes simplex virus type 1 (HSV-1) establishes latency in human senso ry ganglia, during which time the viral genome is transcriptionally si lent with the exception of the latency-associated transcripts (LATs). The most abundant LAT is a 2-kb RNA whose biosynthesis is poorly chara cterized. The 2-kb LAT may be a primary transcript, or its synthesis m ay involve splicing and/or other forms of processing. Two potential RN A polymerase II promoters (LAP1 and LAP2) upstream of the 2-kb LAT 5' end have been identified. To investigate the role played by LAP1 and L AP2 in the synthesis of the 2-kb LAT under lytic and latent conditions , we analyzed HSV-1 mutants which contain deletions of one or both of these promoters. During lyric infection in cell culture, the cis eleme nts critical for the normal accumulation of the 2-kb LAT were mapped t o LAP2, while LAP1 sequences were largely dispensable. The 5' ends of the major 2-kb LATs produced by the wild-type and LAP deletion viruses were examined by primer extension analysis and were all found to be i dentical (+/-2 bp). The accumulation of the 2-kb LAT during latent inf ections of murine trigeminal ganglia was examined by Northern (RNA) bl ot and by reverse transcription-PCR. In contrast to the results found in lytic infections, the critical cia elements needed for 2-kb LAT acc umulation during latency were mapped to LAP1. Deletion of LAP1 resulte d in a 500-fold reduction in 2-kb LAT accumulation, whereas deletion o f LAP2 resulted in only a 2- to 3-fold reduction. Deletion of both LAP 1 and LAP2 resulted in undetectable levels of the 2-kb LAT. Our result s indicate that both LAP1 and LAP2 are critical for 2-kb LAT expressio n but under different conditions. LAP1 is essential for LAT expression during latency, while LAP2 is primarily responsible for LAT expressio n in lyric infections in cell culture. LAP1 and LAP2 may prove to be f unctionally independent promoter elements that control 2-kb LAT expres sion during different stages of HSV-1 infections.