Figure 3.

IFITMs from mycobacteria inhibit influenza virus infection when expressed in human cells. (A) HEK293T cells were transfected overnight with equal amounts empty vector or vector expressing human HA-IFITM3, HA-MAB IFITM3 or HA-MAV IFITM3. Lysates were analyzed by Western blotting with anti-HA antibody staining and anti-GAPDH staining as a loading control; (B) HEK293T cells were transfected overnight with equal amounts of empty vector, human HA-IFITM3, murine HA-IRGM1, or HA-MAV IFITM, or half the concentration of HA-MAB IFITM. Cell lysates were analyzed by Western blotting with anti-HA antibody staining and anti-GAPDH staining as a loading control; (C,D) Cells transfected as in B were infected with IAV H1N1 strain PR8 at an MOI of 2.5 or IAV H3N2 strain X31 or SeV at an MOI of 5.0. Cells were then collected and stained with anti-HA and anti-IAV nucleoprotein antibodies or anti-SeV antiserum. Cells were analyzed for percent infection by flow cytometry based on anti-IAV nucleoprotein or anti-SeV antibody staining in comparison to non-infected controls. The average percent infection for vector control cells was set to 1 for each infection. At least three independent experiments, each with triplicate measurements, were averaged and graphed. Error bars indicate standard errors of the means. * p < 0.01 compared to vector control by Student’s t test.