SUMOylation protects FASN against proteasomal degradation in breast cancer cells treated with grape leaf extract

Background: Existing therapeutic strategies for breast cancer are limited by tumor recurrence and drug-resistance. Several epidemiological studies indicate that antioxidant plant-derived compounds such as flavonoids reduce adverse outcomes and have been identified as a potential source of antineoplastic agent with less undesirable side effects. Activation of lipid metabolism is an early event in carcinogenesis and a central hallmark in breast cancer. In fact, inhibition of fatty-acid synthesis in breast cancer results in cytotoxicity that triggers apoptosis. Here, we describe the novel regulation of lipid metabolism in breast cancer cells whereby the protein stability and degradation of fatty-acid synthase (FASN), the key enzyme in de novo fatty-acid synthesis, is regulated by SUMOylation.Methods: The phenolic characterization were analyzed by Liquid Chromatography-Mass Spectrometry (LCMS). Profile protein contents was evaluated by Mass Spectrometry (LC-MS/MS). The experiments were performed using MCF7, SKBR-3 human carcinoma cell lines and MCF-12A breast epithelial cell line treated with Vermentino hydroalcoholic extract in dose and time course responses. Protein and mRNA levels were analyzed by western blotting/Co-immunoprecipitation (Co-IP) and RT-PCR, respectively. The number of viable cells and the cell-surviving has been detected by MTT and clonogenicity assays. Apoptotic induction was determined by Flow Cytometric assay using Annexin V-FITC and sorted by A FACSC analysis.Results: We first tested the potential antitumorigenic effects of Vitis vinifera L. cv. Vermentino leaf hydroalcoholic extract in MCF-7 and SKBR-3 breast cancer cell lines and found that this compound demonstrated cytotoxic effects. We went on to determine that FASN and UBC9, the sole E2 enzyme required for SUMOylation, were significantly reduced by treatment with the Vermentino extract. Moreover, we found that FASN was SUMOylated in human breast cancer tissues and cell lines. Finally, lack of SUMOylation caused by SUMO2 silencing reduced FASN protein stability.Conclusion: Altogether, these results suggest that SUMOylation protects FASN against proteasomal degradation and may exert oncogenic activity through alteration of lipid metabolism in breast cancer. Importantly, we found that these effects were significantly inhibited by treatment with Vermentino leaf extract, which supports the additional validation of the therapeutic value of this compound.