Introduction. The G protein-coupled estrogen receptor (GPER) is known for its ability to mediate rapid estrogen signaling in diverse normal and malignant cell contexts, including breast cancer (BC). Of note, the role of the GPER in promoting pro-tumorigenic traits in triple-negative breast cancer (TNBC) has been suggested. In this study, we sought to provide novel insights into the transcriptionally guided biological behavior of TNBC cells lacking GPER expression.
Methods. GPER knock-out (KO) MDA-MB-231 TNBC cells were obtained using the CRISPR/Cas9 genome editing technology. RNA sequencing (RNA-seq) and Gene Ontology (GO) enrichment analyses served to assess the differentially expressed genes (DEGs) between the GPER KO and wild-type (WT) cells and their biological role. Chromatin immunoprecipitation assays, real-time PCR, immunoblots, immunofluorescence, ELISA, and flow cytometric experiments, as well as the use of RNA interference techniques, allowed us to uncover the molecular routes implicated in the biological features triggered by GPER silencing in the TNBC cells. Survival analyses were performed on TNBC patients ffrom the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC) dataset.
Results. The RNA-seq data revealed that GPER silencing in MDA-MB-231 cells induces a pro-apoptotic gene signature. Accordingly, the biological assays demonstrated that GPER KO cells exhibit enhanced mitochondria-dependent apoptotic death with respect to these levels in WT cells. Mechanistically, we found that reduced cAMP levels in GPER KO cells may trigger the activation of the pro-apoptotic JNK/c-Jun/p53/Noxa pathway. In accordance with these results, the bioinformatics analyses on the TNBC patients revealed that high NOXA gene expression levels are associated with better outcomes.
Conclusions. Collectively, our findings unveil the role of the GPER in sustaining anti-apoptotic signals in TNBC cells, thus suggesting this receptor as a potential valuable therapeutic target for preventing the progression of this malignancy.
