Figure 2

ERα expression enhances autophagic flux and differentially modulates key autophagy pathway-related protein expression. (a and b) Protein extracts from vehicle or BafA₁ treated SK-01, SK-ERα, SK-ERβ and MCF-7 cells were subjected to western blot analysis and calculation of autophagic flux using anti-LC3 and NBR1 antibodies. Tubulin was used as a loading control. (a) ERα expression of the different cell lines was shown via western blot analysis and autophagic flux was determined by the accumulation of LC3-II in a 6 h treatment period with 500 nM BafA₁. Therefore, normalized LC3-II levels in the absence of the lysosomal inhibitor were subtracted from the corresponding levels obtained in the presence of BafA₁. (b) Cells were treated as in (a) and autophagic flux was calculated by NBR1 expression determined by western blot. (c–h) Western blot analysis of protein extracts from untreated cells were performed for detection of indicated proteins. In the diagrams (lower panel each) levels of proteins are depicted after normalization to corresponding Tubulin levels. Values of three independent experiments in each panel are expressed as mean±S.E.M. and control SK-01 cells were set to 100%. (*) on bars represents statistical significance of P<0.05 comparing two groups and n.s. displays no statistical significant difference