c-Src Modulates Estrogen-Induced Stress and Apoptosis in Estrogen-Deprived Breast Cancer Cells

The emergence of anti-estrogen resistance in breast cancer is an important clinical phenomenon affecting long-term survival in this disease. Identifying factors that convey cell survival in this setting may guide improvements in treatment. Estrogen (E-2) can induce apoptosis in breast cancer cells that have been selected for survival after E-2 deprivation for long periods (MCF-7:5C cells), but the mechanisms underlying E-2-induced stress in this setting have not been elucidated. Here, we report that the c-Src kinase functions as a key adapter protein for the estrogen receptor (ER, ESR1) in its activation of stress responses induced by E-2 in MCF-7:5C cells. E-2 elevated phosphorylation of c-Src, which was blocked by 4-hydroxytamoxifen (4-OHT), suggesting that E-2 activated c-Src through the ER. We found that E-2 activated the sensors of the unfolded protein response (UPR), IRE1a (ERN1) and PERK kinase (EIF2AK3), the latter of which phosphorylates eukaryotic translation initiation factor-2 alpha (eIF2 alpha). E-2 also dramatically increased reactive oxygen species production and upregulated expression of heme oxygenase HO-1 (HMOX1), an indicator of oxidative stress, along with the central energy sensor kinase AMPK (PRKAA2). Pharmacologic or RNA interference-mediated inhibition of c-Src abolished the phosphorylation of eIF2 alpha and AMPK, blocked E-2-induced ROS production, and inhibited E-2-induced apoptosis. Together, our results establish that c-Src kinase mediates stresses generated by E-2 in long-term E-2-deprived cells that trigger apoptosis. This work offers a mechanistic rationale for a new approach in the treatment of endocrine-resistant breast cancer.