FIG. 8.
Immunoprecipitation of IE62 protein from melanoma cells infected with IE63 mutant viruses with anti-IE63 antiserum and expression of VZV immediate-early, early, and late proteins by IE63 mutant viruses. As shown in panel A, IE62 in lysates of infected cells was immunoprecipitated with monoclonal antibody against IE62. IE63 that coimmunoprecipitated with IE62 was detected with polyclonal rabbit antiserum against IE63 (even-numbered lanes) or nonspecific murine IgG (odd-numbered lanes). Samples tested included lanes 1 and 2, uninfected melanoma cells; lanes 3 and 4; rOKA/ORF63rev-infected cells; lanes 5 and 6, rOKA/ORF63rev[T171]-infected cells; lanes 7 and 8, rOKA/ORF63rev[S181]-infected cells; and lanes 9 and 10, rOKA/ORF63rev[S185]-infected cells. As shown in panel B, melanoma cells were infected with 5 × 104 PFU of the test virus and harvested at 4 days postinfection. Cell lysates were evaluated for expression of immediate-early proteins IE62 and IE63 and ORF4 proteins, the ORF47 viral kinase, which is an early protein, and the late protein gE, with rabbit polyclonal antiserum specific for IE62, IE63, ORF4, or ORF47, and a murine monoclonal antibody to gE. The specimens tested were lane 1, rOka (contains ORFs 63 and 70); lane 2, rOKA/ORF63rev (one copy of ORF63 at a nonnative site); lane 3, rOKA/ORF63rev[T171]; lane 4, rOKA/ORF63rev[S181]; and lane 5, rOKA/ORF63rev[S185].