Figure 9.

Processing IAV HA. (A) 293T cells (1 × 10⁵) plated in a 24-well plate were inoculated with IAV (MOI = 0.001, 0.01). Cell lysates were prepared after the indicated periods and subjected to electrophoresis and western blotting with an anti-IAV HA antibody. (B) IAV HA₀ was prepared from IAV-infected U937 lymphoma cells (MOI = 0.1), which permit IAV proliferation, and was used in processing experiments in vitro. IAV HA₀ lysates were incubated for 30 min at room temperature with TPCK-treated trypsin (0, 3, or 9 μg/ml) or with the indicated lysates expressing PRSS3, EK, EK-X2, or vector only (–), and then subjected to western blotting with an anti-IAV HA antibody. The 65-kDa precursor IAV HA₀ and the cleaved 25-kDa C-terminal IAV HA₂ fragment are indicated by arrows. The HA₀ and HA₂ intensities and the HA₂/HA₀ ratio are shown below.