THE PRODUCT OF ORF-O LOCATED WITHIN THE DOMAIN OF HERPES-SIMPLEX VIRUS-1 GENOME TRANSCRIBED DURING LATENT INFECTION BINDS TO AND INHIBITS IN-VITRO BINDING OF INFECTED CELL PROTEIN-4 TO ITS COGNATE DNA SITE

The partially overlapping ORF P and ORF O are located within the domai ns of the herpes simplex virus 1 genome transcribed during latency, Ea rlier studies have shown that ORF P is repressed by infected cell prot ein 4 (ICP4), the major viral regulatory protein, binding to its cogna te site at the transcription initiation site of ORF P. The ORF P prote in binds to p32, a component of the ASF/SF2 alternate splicing factors ; in cells infected with a recombinant virus in which ORF P was derepr essed there was a significant decrease in the expression of products o f key regulatory genes containing introns, We report that (i) the expr ession of ORF O is repressed during productive infection by the same m echanism as that determining the expression of ORF P; (ii) in cells in fected at the nonpermissive temperature for ICP4, ORF O protein is mad e in significantly lower amounts than the ORF P protein; (iii) the res ults of insertion of a sequence encoding 20 amino acids between the pu tative initiator methionine codons of ORF O and ORF P suggest that ORF O initiates at the methionine codon of ORF P and that the synthesis o f ORF O results from frameshift or editing of its RNA; and (iv) glutat hione S-transferase-ORF O fusion protein bound specifically ICP4 and p recluded its binding to its cognate site on DNA in vitro, These and ea rlier results indicate that ORF P and ORF O together have the capacity to reduce the synthesis or block the expression of regulatory protein s essential for viral replication in productive infection.