IDENTIFICATION OF A NOVEL LATENCY-SPECIFIC SPLICE DONOR SIGNAL WITHIN THE HERPES-SIMPLEX VIRUS TYPE-1 2.0-KILOBASE LATENCY-ASSOCIATED TRANSCRIPT (LAT) - TRANSLATION INHIBITION OF LAT OPEN READING FRAMES BY THE INTRON WITHIN THE 2.0-KILOBASE LAT

Herpes simplex virus type 1 establishes latent infection in trigeminal ganglia of mice infected via the eye. A family of three collinear viral transcripts (LATs), 2.0, 1.5, and 1.45 kb, is present in latently infected ganglia. To characterize these LATs, lambda-gt10 cDNA libraries were constructed with RNAs isolated from the trigeminal ganglia of latently infected mice. A series of recombinant bacteriophage were isolated containing cDNA inserts covering 1.7 kb of the 2.0-kb LAT. Splice junctions of the smaller LATs and the 3' end of the 2.0-kb LAT were identified by sequence analysis of RNA polymerase chain reaction products. No splice was detected in the 2.0-kb LAT. The 3' end of the 2.0-kb LAT in vivo is upstream of a consensus splice acceptor site, which does not support the hypotheses that the 2.0-kb LAT is an intron. However, the data are consistent with the possibility of a short leader sequence or multiple LAT transcription start sites. To generate the smaller 1.5- and 1.45-kb LATs, there is a 559-nucleotide intron spliced from the 2.0-kb LAT in strain F and a 556-nucleotide intron in strain 17+. The nucleotide sequences at the 5' and 3' ends of these introns are characteristic of spliced transcripts from eukaryotic protein-encoding genes, with one significant difference; i.e., the 5' end of the LAT intron is GC instead of the consensus sequence GT. This splice donor sequence is conserved in herpes simplex virus type 1 strains F, 17+, and KOS. Processing of the 2.0-kb LAT to form the spliced LATs preserves two open reading frames (ORFs) at the 3' end of the LAT; no new ORFs are created. Splicing of the LATs positions a 276-nucleotide leader sequence close to these ORFs and removes an intron that inhibits their translation in vitro. The novel 5' structure of the intron within the 2.0-kb LAT may be part of a control mechanism for transcription processing that results in splicing of the LATs only in sensory neurons during latent infection and reactivation but not during the viral replication cycle.