A 348-BASE-PAIR REGION IN THE LATENCY-ASSOCIATED TRANSCRIPT FACILITATES HERPES-SIMPLEX VIRUS TYPE-1 REACTIVATION

Latency-associated transcript (LAT) promoter deletion mutants of herpe s simplex virus type 1 have a reduced capacity to reactivate following adrenergic induction in the rabbit eye model. We have mapped a reacti vation phenotype within LAT and describe the construction of recombina nts in which poly(A) addition sites have been placed at intervals with in the LAT region to form truncated LAT transcripts, These mutants loc alize the induced reactivation phenotype to the 5' end of LAT. To furt her define this region, we constructed a recombinant containing a 348- bp deletion located 217 bp downstream of the transcription start site of the 8.5-kb LAT. This virus, 17 Delta 348, expresses LAT but exhibit s a significantly reduced ability to reactivate following epinephrine iontophoresis into the cornea. Quantitative DNA PCR analysis reveals t hat 17 Delta 348 establishes a latent infection within rabbit trigemin al ganglia with the same efficiency as does either the rescuant or wil d-type virus. The region deleted in 17 Delta 348 encodes three potenti al translational initiators (ATGs) which we have mutated and demonstra ted to be dispensable for epinephrine-induced reactivation. In additio n, three smaller deletions within this region have been constructed an d were shown to reactivate at wild-type (parent) frequencies. These st udies indicate that an undefined portion of the 348-bp region is requi red to facilitate induced reactivation. Sequence analysis of this 348- bp region revealed a CpG island which extends into the LAT promoter an d which possesses homology to conserved elements within the mouse and human XIST transcript encoded on the X chromosome. Possible implicatio ns of these elements in the regulation of LAT expression are discussed .