Fig. 6. Cellular distribution of overexpressed TACE. Lysates of COS-7 cells overexpressing haTACE were treated in vitro with PNGase F or Endo H as described in Materials and Methods and submitted to western blot analysis using the anti-HA epitope antibody to detect overexpressed haTACE (A) or the cytoplasmic-domain-specific TACE antibody to detect endogenous TACE (B). Mock (lanes 1 and 3) samples without PNGase F or EndoH. Samples treated with PNGase F (lanes 2) or with EndoH (lanes 4). Immature and mature forms of endogenous TACE are indicated by i and m, respectively. Overexpression of haTACE was analyzed by flow cytometry at the cell surface (C) or intracellularly (D) as described in Materials and Methods. Solid lines (unshaded areas) depict untransfected cells, and shaded areas haTACE transfected cells.