ROLE OF HERPES-SIMPLEX VIRUS TYPE-1 UL46 AND UL47 IN ALPHA-TIF-MEDIATED TRANSCRIPTIONAL INDUCTION - CHARACTERIZATION OF 3 VIRAL DELETION MUTANTS

The transcriptional induction of the alpha or immediate-early gene class of herpes simplex virus type 1 effected by the alpha trans-induction factor (alpha-TIF, ICP25, VP16, Vmw65) requires an alpha-specific cis-acting site. Increased transcription does not result from the direct, independent binding of alpha-TIF, but rather from an alpha-TIF-dependent formation of a protein-DNA complex containing, in addition to alpha-TIF, at least one host cell factor. One of the host factors is a POU domain protein which recognizes an octamer element in the alpha-specific consensus. There is evidence that alpha-TIF may drive the formation of multiple protein-DNA complexes containing a POU protein and additional host factors. Previously, the gene products of UL46 and UL47 have been implicated in modulating the alpha-TIF-dependent transcriptional induction of alpha genes. Our current studies have extended these analyses from a transient-expression system to a series of viral deletion mutants. In these studies we demonstrate that neither UL46- nor UL47-encoded gene product, either separately or in combination, is required for viral growth in cell culture. The absence of UL47 reduces by up to 80% the ability of the virus to induce an alpha-regulated thymidine kinase reporter gene resident in 143TK- cells. Autoradiograms of [S-35]methionine pulse-labeled infected cell proteins, separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, show that deleting UL46 and/or UL47 has no discernable effect on the synthesis of alpha-TIF or alpha-TIF-containing proteins. Subsequent Western immunoblot analysis, with rabbit anti-alpha-TIF antibodies made to an alpha-TIF-Staphylococcus aureus protein A fusion, demonstrated that the accumulation and steady-state levels of alpha-TIF or alpha-TIF-containing proteins was indistinguishable from that of the thymidine kinase-negative isogenic parental virus, R-DELTA-305.