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The effect of Sirt3 expression on human breast cancer cells in normoxia and hyperoxia

Sirtuin 3 (Sirt3), a major mitochondrial NAD+ dependent deacetylase, has bifunctional role in cancer tumorigenesis, acting as both oncogene and tumor suppressor, depending on the tissue and cancer-type specific metabolic programs. Changes in its expression are associated with the excessive production of reactive oxygen species (ROS), thus contributing to mitochondrial dysfunction and age-related pathologies. Hyperoxic treatment (i.e. generator of ROS) was shown to support some tumorigenic properties, but finally suppresses growth of certain mammary carcinoma cells. 70% of all breast cancer cases are estrogen receptor (ER) positive and express ER-alpha (ER- α). While ER-α positive cancers are more receptive to hormonal therapy, triple negative breast cancers (TNBC) are characterized by an aggressive behaviour and the lack of targeted therapeutic strategies. Due to strikingly reduced Sirt3 level in many breast cancer cells, we aimed at deciphering the effect of de novo Sirt3 expression in normoxic and hyperoxic conditions in the human breast cancer cells. Although we have recently shown that Sirt3 acts as a tumor suppressor in non-invasive (ER-α positive) breast cancer cells, it remains unclear whether this effect is mediated through ER-α signalling pathway. Therefore, we stably transfected both MCF-7 (ER-α positive) and MDA-MB-231 (TNBC) cells with Flag-tagged Sirt-3 plasmid and characterized Sirt-3 overexpressing clones in normoxic and hyperoxic conditions. To further characterize clones and decipher the cause of the observed differences in their response, we used combination of treatments to alter the expression and activity of different proteins. We monitored the expression of proteins involved in mitochondrial biogenesis, glycolysis, metabolic regulation and antioxidant defense. Furthermore, we compared the growth rate, metabolic activity, mitochondrial ROS production and the cell cycle of the clones. Initial findings showed enhanced susceptibility of MCF-7 cells to hyperoxia and decreased cellular growth upon de novo Sirt3 expression. On the other hand, Sirt-3 markedly promoted growth of highly invasive MDA-MB-231 cells. Collectively, results suggested that Sirt-3 may either have a tumor suppressing or tumor promoting role in breast cancer cells depending on their invasiveness, thus giving us a rationale for further studies on Sirt3 and hyperoxia as an adjuvant tumor therapy in breast cancer malignancies.

Hyperoxia ; MCF-7 ; ROS ; sirtuin 3

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