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. 1998 Jun 1;141(5):1255–1266. doi: 10.1083/jcb.141.5.1255

Figure 8.

Figure 8

(a) FADD and FLICE coimmunoprecipitate with Fas in the presence of E1B 19K. HeLa cells were transfected with 7dl vector control (lanes 1 and 2), FADD, FLICE, and CrmA (lanes 3 and 4), or FADD, FLICE, E1B 19K, and CrmA (lanes 5 and 6) and then immunoprecipitated with an anti–APO-1 antibody 48 h posttransfection. In lanes 1, 3, and 5, cells were first stimulated with anti–APO-1 antibody, lysed, and then immunoprecipitated with the addition of protein A–Sepharose beads. In lanes 2, 4, and 6, cells were lysed first and then immunoprecipitated with anti–APO-1 antibody. Lanes 7–9 correspond to whole cell extracts used to verify equal protein expression. Samples were resolved on a 15% SDS-PAGE and subjected to Western analysis with an anti-FADD polyclonal antibody (top panel) and an anti-HA polyclonal antibody (bottom panel). (b) E1B 19K blocks upstream of FLICE activation. HeLa cells were transfected with 7dl vector control, FLICE, or FLICE plus FADD alone or in the presence of E1B 19K at concentrations indicated in the viability assay (refer to Materials and Methods). Whole cell extracts were made 48 h posttransfection, resolved by SDS-PAGE, and then immunoblotted with an anti-HA polyclonal antibody.