Apoptosis induction by FnE requires internalization by the Aα52-83 motif. (A) Representative FACS analysis of FITC-labeled FnE uptake (2 μM) by JEG3 cells in suspension. Uptake only occurs at 24°C in the presence of Ca2+. (B) Uptake of FITC-labeled FnE into JEG3 cells is competitively inhibited by a 100-fold excess of unlabeled FnE and Aα-chain fragments containing the 52-83 motif and by intact fibrinogen, but not by RGD-containing peptides. (C-G) JEG3 cells in suspension culture were incubated with FITC- or FAM-labeled preparations of the indicated peptides (2 μM, 24°C). Distribution of fluorescence was determined by confocal microscopy of cytospin preparations. (H-M) Representative TUNEL staining of adherent JEG3 cells cultured for 48 hours in the presence of 2 μM 17-83 peptide (H), 2 μM 17-83 peptide plus a 100-fold excess (200 μM) 52-83 peptide; 20 μM 17-37– (K), 17-37-TAT– (L), or 17G/A-37-TAT–peptides (M). Bars represent 50 μm (C-I) or 200 μm (K-M).
