Figure 1. Phenotypic impact of AIB1 depletion on estradiol (E2) growth response in MCF-7 or MCF-7:5C cells.

(A) The experimental paradigm. The differential responses to estradiol (E2) treatment of MCF-7 (cell growth) and long-term estrogen deprived MCF-7:5C cells (apoptosis) are indicated. Proteomics profiles of the two cell lines at baseline and after a brief (2 h) E2 treatment were generated using immunoprecipitations (IP). Proteins interacting with AIB1 or phosphotyrosine containing protein complexes were isolated by IP followed by mass spectrometry. Data were then subjected to an integrated bioinformatics analysis of signaling pathways and protein networks. (B,C) Reversal of E2-dependent effects on MCF-7 and MCF-7:5C after depletion of endogenous AIB1 protein using two different lentiviral shRNAs. MCF-7 or MCF-7:5C cells were infected with lentiviral particles expressing control or AIB1-targeting shRNAs. (B) RNAi-mediated knockdown was assayed by Western blot analysis for AIB1 relative to an actin loading control. (C) Cell growth was assayed 6 days after plating without or with E2. The E2 effect is shown relative to the respective untreated controls (mean ±S.E.M.). Closed circles: control shRNA; Open circles (red): AIB1 shRNA. #, p<0.05 E2 treatment effect vs. no treatment in control shRNA cells; *, p<0.05 E2 treatment effect in control shRNA cells vs. E2 treatment in AIB1 depleted cells. Representative data from one of at least three independent experiments are shown.