Figure 4
From: REP1 inhibits FOXO3-mediated apoptosis to promote cancer cell survival
REP1 interacts with FOXO3. (a and b) Positive protein–protein interactions were determined by monitoring cell growth over 3 days on a medium lacking leucine (a), and by the formation of blue colonies on X-gal plates containing galactose at 30 °C (b). The values of β-galactosidase activity (unit) estimated by adding o-nitrophenyl β-D-galactopyranoside (ONPG) are indicated below their corresponding lanes. Graph represents two independent experiments performed in triplicate. Error bars represent S.D. from the mean. **P<0.001. (c) Co-immunoprecipitation of REP1 and FOXO3. HEK293 cells were co-transfected with the indicated constructs. Lysates of the transfected cells immunoprecipitated with anti-HA antibody (left) or anti-FLAG antibody (middle), followed by western blotting using anti-FOXO3 and anti-REP1 antibodies. The input represents 5% of whole-cell lysates (WCL) used in immunoprecipitation (right). (d–e) Endogenous REP1 binds to endogenous FOXO3 in HCT116 cells. Immunoprecipitation was performed using rabbit IgG, anti-FOXO3 (d) or anti-REP1 (e), followed by western blotting using the indicated antibodies
