FIG. 2.

rM51R-M virus infection induces the movement of the Fas death receptor to the cell surface. (A) L929 cells were mock infected or infected with rWT or rM51R-M virus. At the indicated times postinfection, cell lysates were generated and analyzed via Western blotting with antibodies for the Fas death receptor and β-actin. (B) Western blots were quantitated via densitometric analysis. Levels of Fas protein expression were normalized to levels of actin for each sample. Results represent the averages of three independent experiments ± standard errors of the means (SEM). (*P < 0.05 relative to mock-infected samples). (C) L929 cells were mock infected (green line), infected with rWT virus (blue line) or rM51R-M virus (red line), or treated with etoposide (orange line). At 24 h postinfection or posttreatment, levels of Fas surface expression were analyzed via flow cytometry using anti-Fas antibodies conjugated to PE. Background was determined by using a PE-conjugated mouse immunoglobulin G2 isotype control antibody. (D) Fas surface expression was analyzed at the indicated times postinfection by quantitating flow cytometry data. Data are expressed as the changes (n-fold) above mock-infected cells. Results are from three independent experiments ± SEM. (E) To measure the effect of blocking FasL signaling, L929 cells were untreated, infected with rM51R-M virus, or treated with rFasL and cycloheximide (CHX) in the presence or absence of FasL neutralizing antibody (Nok1). Apoptosis was measured via flow cytometry by staining with annexin V and 7-AAD. Data are presented as percentages of annexin V-positive cells and represent the average ± SEM of three experiments. hpi, hours postinfection.