Figure 4.

Cyr61 secretion in breast cancer cells. (A) Detection of Cyr61 secretion in MDA-MB-231 (MDA-231) and BC-M1 by Western blot analysis. Cells were treated with Brefeldin A to block the protein secretion. The supernatant of the cell culture medium contains the secreted protein fraction, whereas the pellet contains the detached cells. As the total protein amount for two of the three biological replicates of the MDA-231 detached cells (untreated) was below 50 µg, a Western blot lacking this sample is shown. The 78 kDa glucose-regulated protein (Grp78) was analysed as the positive control for a cellular response to Brefeldin A treatment. (B) A comparison of the protein levels in the parental cell line MDA-231 and its bone metastatic subline MDA-231 B02 in the non-stimulated state by Western blot analysis. (C) The activation of the cells (detected as pAKT S473) upon EGF stimulation, and the analysis of the cytoplasmic Cyr61 levels by Western blot analysis. (D) The secreted Cyr61 levels in the cell culture supernatants after EGF stimulation. For a statistical analysis of the values, see Tables S2 and S3. The corresponding scatterplots are shown in Figure S7.