THE HERPES-SIMPLEX VIRUS-1 U(L)15 GENE ENCODES 2 PROTEINS AND IS REQUIRED FOR CLEAVAGE OF GENOMIC VIRAL-DNA

Previous studies have shown that a ts mutant [herpes simplex virus 1(m P)ts66.4] in the U(L)15 gene fails to package viral DNA into capsids ( A. P. W. Poon and B. Roizman, J. Virol. 67:4497-4503, 1993) and that a lthough the intron separating the first and second exons of the U(L)15 gene contains U(L)16 and U(L)17 open reading frames, replacement of t he first exon with a cDNA copy of the entire gene does not affect vira l replication (J. D. Baines, and B. Roizman, J. Virol. 66:5621-5626, 1 992). We report that (i) a polyclonal rabbit antiserum generated again st a chimeric protein consisting of the bacterial maltose-binding prot ein fused in frame to the majority of sequences contained in the secon d exon of the U(L)15 gene reacted with two proteins with M(r) of 35,00 0 and 75,000, respectively, in cells infected with a virus containing the authentic gene yielding a spliced mRNA or with a virus in which th e authentic U(L)15 gene was replaced with a cDNA copy. (ii) Insertion of 20 additional codons into the C terminus of U(L)15 exon II caused a reduction in the electrophoretic mobility of both the apparently 35,0 00- and 75,000-M(r) proteins, unambiguously demonstrating that both sh are the carboxyl terminus of the U(L)15 exon II. (iii) Accumulation of the 35,000-M(r) protein was reduced in cells infected and maintained in the presence of phosphonoacetate, an inhibitor of viral DNA synthes is. (iv) The U(L)15 proteins were localized in the perinuclear space a t 6 h after infection and largely in the nucleus at 12 h after infecti on. (v) Viral DNA accumulating in cells infected with herpes simplex v irus 1(mP)ts66.4 and maintained at the nonpermissive temperature was i n an endless (concatemeric) form, and therefore U(L)15 is required for the cleavage of mature, unit-length molecules for packaging into caps ids.