Figure 10. GOLPH3 overexpression in breast cancer cells confers mTOR activation and autophagy resistance. We overexpressed GOLPH3 in MDA-MB-231 breast cancer cells, to examine its functional effects on mTOR activation and autophagy resistance. (A) Note that GOLPH3 was successfully stably expressed in MDA-MB-231 cells, as detected using antibodies directed against the HA-epitope tag. (B) Under basal cell culture conditions, GOLPH3 activates mTOR hyper-phosphorylation, and a loss of cathepsin B expression. (C) Under conditions of starvation [4 h in HBSS (40 mM HEPES, pH 7.2, in Hank’s Balanced Salt Solution)], GOLPH3 confers autophagy resistance, as evidenced by a loss of BNIP3 and LC3-I/II expression. In panels A-C, immunoblotting with β-actin is shown as a control for equal protein loading.