FIGURE 1.

RIG-I, NIK, and IKKα mediate RSV-induced p52 formation. A, A549 cells were treated with LIGHT at 0, 0.5, 1, 3, and 6 h, as well as infected with RSV (multiplicity of infection 1) at 12 and 24 h. Whole cell extracts were collected and Western immunoblot was conducted to detect the expression of p100 and its proteolytic product p52. β-Actin staining was used as a loading control. B, A549 cells were treated with RSV, UV-inactivated RSV, or RSV conditioned medium (CM) for the indicated times (in h, bottom). Right panel, cells were RSV infected and treated with cycloheximide (CHX, 25 μg/ml) for the indicated times. The abundance of p100 and p52 were determined by Western immunoblot. β-Actin staining was used as a loading control. C, A549 cells transfected with control (Con) siRNA and RIG-I siRNA for 48 h were then RSV infected for 0 and 24 h. Total RNA was extracted and assayed by RT-PCR to measure the expression of RIG-I (upper panel). β-Actin is a control. Shown is an ethidium bromide-stained agarose gel. Whole cell lysates were prepared from the same cell treatment and p100/p52 by Western immunoblot using NH₂-terminal anti-NF-κB2 Ab (lower panel). The blot was probed with β-actin as a loading control. D, WT, Nik^-/-, Ikkα^-/-, and Ikkγ^-/- MEFs were infected by RSV (multiplicity of infection 1) for 0 and 24 h. Whole cell lysates were assayed by Western immunoblot to detect p100 expression and p52 formation (indicated by asterisk). β-Actin was a loading control. p52 was detected only in RSV-infected WT and Ikkγ^-/- cells.