FIG. 3.

Complementation of the HSV-1-induced SINrep/LacZ replicon by a defective helper expressing the Sindbis virus structural proteins. (A) Induction of β-galactosidase activity in V2-33, V2-34, and V3-45 Vero cell lines was dependent on infection with HSV-1. Three independently transformed cell clones, V2-33, V2-34, and V3-45, were plated into the wells of a 24-well dish. Subconfluent monolayers (50% confluence) of the cells were transfected with the defective helper cDNA plasmid, p987DHBBneo. After 72 h, the cells were either mock infected or infected with HSV-1 at an MOI of 5 in the presence of acyclovir. At 30 h postinfection, cell extracts (10 μl) were assayed for β-galactosidase activity as described in Materials and Methods. (B) Production of SINrep/LacZ particles was dependent on HSV-1 infection of ICP8SINrep/LacZ-transformed Vero cell lines (V2-33, V2-34, and V3-45). A sample (200 μl) of the medium harvested from the Vero cell lines (assayed as described for panel A) was inoculated onto BHK cells in the wells of a 24-well dish. At 18 h postinfection of the BHK cells, lysis buffer (250 μl) was added to the cells and the extracts (50 μl) were assayed for β-galactosidase activity. (C) Production of SINrep/LacZ particles was dependent on transfection of the Vero cell lines with the defective helper plasmid. Subclones of V2-33 and V2-34 cells were infected with HSV-1 (MOI = 10). At 5 h postinfection the cells were either transfected with the helper plasmid p987DHBBneo or mock transfected. The media were collected 18 h later and inoculated onto BHK cells, and 18 h later β-galactosidase activity was assayed as described for panel B.