Fig. 5. Bcl-2 over expression modifies the level of junctional proteins and decreases the surface E-cad. (A) MCF-7 and MCF-7B cells were grown to confluence in normal medium (day 0). Replicate cultures were transferred into estrogen-free medium for 2 to 16 days. Equal amounts of TCEs from control and estrogen-depleted cultures was resolved on 10% (Bcl-2 and tubulin) or 6% (E-cad, -cat, ß-cat, Pg, p120 and ZO-1) gels and processed for IB with various antibodies at concentrations listed in Table 1. (B) MCF-7 and MCF-7B were grown in normal medium and duplicate cultures were transferred into estrogen-deprived media for 12 days. Cultures were biotinylated as described in Materials and Methods and TCEs prepared. From each line, 250 µg of TCE was precipitated with streptavidin-agarose beads. Biotinlylated proteins were eluted from the beads and, together with 50 µg of TCEs, processed for IB with E-cad antibodies. To verify comparable loading the TCE blot was reprobed for tubulin. (C) Confluent MCF-7 and MCF-7B cells were either fixed and then permeabilized (Total) or CSK-extracted and then fixed (Surface) and processed for staining with an E-cad antibody (Transduction Laboratories). Scale bar: 50 µm.