Figure 3.

Nuclear translocation of CrkI, CrkII, and CrkL by SH3 binding-competent NS1 proteins demonstrated by subcellular fractionation. (A) Western blot analysis of cytoplasmic (C) and nuclear extracts (N) prepared from A549 cells that were mock-infected (MOCK) or infected for 24 h with recombinant A/WSN containing either the wild-type (WT) or the K217E mutant NS1 from A/Mallard virus at an MOI 2. In addition to antibodies against the Crk-family proteins and NS1, the blotted A549 fractions were also probed with antibodies against Histone H3 and α-tubulin to confirm successful separation of nuclear and cytoplasmic fractions. In addition, unfractionated whole cell extracts (WCE) of the infected cells were Western blotted with anti-NS1 and anti-NP antibodies to confirm uniform infection of cells by the different viruses; (B) Same as (A) except that the cells were infected with recombinant A/WSN virus carrying wild-type (WT) or the T215P mutant NS1 from A/WSN.