title: Two herpes simplex virus type 1 latency-active promoters differ in their contributions to latency-associated transcript expression during lytic and latent infections.
abstract: Herpes simplex virus type 1 (HSV-1) establishes latency in human sensory ganglia, during which time the viral genome is transcriptionally silent with the exception of the latency-associated transcripts (LATs). The most abundant LAT is a 2-kb RNA whose biosynthesis is poorly characterized. The 2-kb LAT may be a primary transcript, or its synthesis may involve splicing and/or other forms of processing. Two potential RNA 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 LAP2 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 lytic infection in cell culture, the cis elements critical for the normal accumulation of the 2-kb LAT were mapped to 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 identical (+/- 2 bp). The accumulation of the 2-kb LAT during latent infections of murine trigeminal ganglia was examined by Northern (RNA) blot and by reverse transcription-PCR. In contrast to the results found in lytic infections, the critical cis elements needed for 2-kb LAT accumulation during latency were mapped to LAP1. Deletion of LAP1 resulted in a 500-fold reduction in 2-kb LAT accumulation, whereas deletion of LAP2 resulted in only a 2- to 3-fold reduction. Deletion of both LAP1 and LAP2 resulted in undetectable levels of the 2-kb LAT. Our results indicate that both LAP1 and LAP2 are critical for 2-kb LAT expression but under different conditions. LAP1 is essential for LAT expression during latency, while LAP2 is primarily responsible for LAT expression in lytic infections in cell culture. LAP1 and LAP2 may prove to be functionally independent promoter elements that control 2-kb LAT expression during different stages of HSV-1 infections.
mesh_terms: D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D002522:Chlorocebus aethiops; D017931:DNA Primers; D017950:Ganglia, Sensory / Q000821:virology; D015967:Gene Expression Regulation, Viral; D018259:Herpesvirus 1, Human / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009447:Neuroblastoma; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000096:biosynthesis / Q000235:genetics*; D012367:RNA, Viral / Q000032:analysis / Q000096:biosynthesis; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D017384:Sequence Deletion; D014158:Transcription, Genetic; D014709:Vero Cells; D017735:Virus Latency
chemical_list: D017931:DNA Primers; D012367:RNA, Viral; D012319:RNA Polymerase II