Figure 5
The inhibitory effect of Ergosterol on protein synthesis and proteasome degradation of breast cancer cells. (a) The protein synthesis assay using Click-iT™ HPG Alexa Fluor™ 594 Protein Synthesis Assay Kit was measured by the CellInsight™ CX7 High Content Screening (HCS) Platform (Thermo Fisher Scientific, Waltham, MA, USA). The mean fluorescence signal was analyzed after being treated with DMSO, ER 20 μM, or cycloheximide 10 μM into MCF-7 and MDA-MB-231 for 24 h. (b) Western blot analysis of the phosphorylation form of 4E-BP1, an initiator of protein synthesis, and (c) the ubiquitin–proteasome system proteins (USP47 and USP9x) after being treated with ER (2 and 20 μM) for 24 h. Data were expressed as mean ± S.D. of three independent experiments. The statistical significance was analyzed and compared to the DMSO control by One-way ANOVA, and Dunnett’s post-hoc analysis with P-value (P < 0.05 *; P < 0.01 **) was considered significant. The original images for western blot analysis were shown in “Supplementary information”.
