Figure 1: Proteomic discovery linking RARA to tamoxifen resistance.
From: Retinoic acid receptor alpha is associated with tamoxifen resistance in breast cancer
(a) Proteomic workflow. The tamoxifen-sensitive MCF7 cell line and the isogenic tamoxifen-resistant cell line LCC2 were treated with 4-OHT for 3 days followed by subcellular fractionation into a DNA-binding (DNAb) and cytosol fraction. Three biological replicates of samples prior and post treatment were digested and iTRAQ-labelled, separated on a narrow-range IPG-IEF strip, pH 3.5–4.5, and analysed by two nLC-MS platforms. (b,c) Ingenuity pathway analysis of changed proteins in the proteomics data indicated a connection of RARA to tamoxifen resistance. (b) Network from comparison of basal levels of MCF7 and LCC2 and (c) changes following 4-OHT treatment in LCC2. Note that ingenuity pathway analysis contains different kinds of data to build connections. For complete networks and explanations, see Supplementary Fig. S4. * indicates proteins in common with e. (d) Western blot of RARA and ER basal protein levels. (***P<0.001, t-test, values represent mean of two experiments in triplicates±s.e.m.). (e) Connection of RA- and E2-regulated genes to proteins in the DNAb proteomics data set. Heatmap of proteins from the DNAb fraction, separated into E2- and RA-regulated genes based on Hua et al.11,23, respectively. Significantly regulated proteins in this study are highlighted with black boxes to the right of the heatmap. The response to E2 and RA is denoted by up or down.
