EFFECT OF THE TRANSCRIPTION START REGION OF THE HERPES-SIMPLEX VIRUS TYPE-1 LATENCY-ASSOCIATED TRANSCRIPT PROMOTER ON EXPRESSION OF PRODUCTIVELY INFECTED NEURONS IN-VIVO

It has been previously reported that the latency-associated transcript (LAT) promoter contains a DNA sequence at the LAT transcription start site which resembles the ICP4 consensus DNA binding site and that thi s site allows ICP4-mediated downregulation of the LAT promoter in tran sient assays (A. H. Batchelor and P. O'Hare, J. Virol. 64:3269-3279, 1 990). We have confirmed these data by showing that an ICP4-expressing plasmid will downregulate lacZ expression from a plasmid containing th e LAT promoter and transcription start site (pJA1) and does not downre gulate lacZ expression from a plasmid in which the start site has been mutagenized (pWAG15). To determine the role of the LAT transcription start site in regulating LAT promoter activity in the context of the v irus, two recombinant viruses, KOS-1 and KOS-15, were studied. KOS-I c ontains an 863-bp portion of the LAT promoter, including the LAT cap s ite, fused to the lacZ gene and inserted into the gC locus (T. P. Marg olis, F. Sedarati, A. T. Dobson, L. T. Feldman, and J. G. Stevens, Vir ology 189:150-160, 1992). The second virus (KOS-15) was constructed in identical fashion, using plasmid pWAG-15, which is not downregulated by ICP4. Vero cells productively infected with KOS-15 produce 10-fold more P-galactosidase than do those infected with KOS-1. In murine dors al root ganglia acutely infected with KOS-1, only 1.2% of dorsal root ganglion neurons that expressed viral antigen also expressed beta-gala ctosidase. In contrast, in KOS-15-infected mice, beta-galactosidase wa s detected in 18% of viral antigen-positive neurons. Similar findings were observed in trigeminal ganglia acutely infected with KOS-1 and KO S-15. Thus, the region encompassing the LAT transcription start site a ppears to play an important role in repression of the LAT promoter act ivity not only in vitro but also in acutely infected neurons in vivo. These results suggest that during productive infection with HSV-1, LAT expression is tightly regulated.