Fig. 6

AGR2 modulated the release of 14-3-3ε and α-actinin 4 upon ER stress and autophagy induction. A, B CMT-U27e or AGR2-KO clones (KO-S10 and KO-S4) were cultured in 1% FBS-containing RMPI and treated with or without 50 nM tunicamycin (denoted Tm) for 14 h. Cell lysate (WCL) samples were collected and subjected to immunoblotting analysis with antibodies specific to indicated proteins. The CM samples were further TCA-precipitated to analyze 14-3-3ε or α-actinin 4 levels with immunoblotting (B). C, D CF41.Mg transfected with pcDNA3.1-HA-AGR2 or the mock control were subsequently cultured in 1% FBS-containing DMED for 14 h with or without addition of 50 nM Tm. Levels of the indicated proteins in WCL (C) and CM (D) were analyzed by immunoblotting. E, F CMT-U27e, KO-S10, or KO-S4 were cultured in serum-free RPMI and treated with 300 nM rapamycin (denoted Rm) or 40 µM chloroquine (denoted CQ) for 16 h. Levels of the indicated proteins in WCL (E) and CM (F) were analyzed by immunoblotting. G, H CF41.Mg transfected with pcDNA3.1-myc.His-AGR2 or the mock control was cultured in serum-free DMEM supplemented with or without 3-MA (2 mM) for 16 h. Levels of the indicated proteins in WCL (G) and CM (H) were analyzed by immunoblotting. The present results were representative data from three independent experiments