NEDDylation negatively regulates ERRß expression to promote breast cancer tumorigenesis and progression.

Estrogen-related receptor beta (ERRß) is downregulated in breast cancer cells and its overexpression in breast cancer patients is positively correlated with an improved prognosis and prolonged relapse-free survival. Here, we unravelled a molecular mechanism for ERRß downregulation in breast cancer. We found that ERRß is a key substrate of the SCF complex and that NEDDylation can activate the Cullin subunits of the SCF complex to target ERRß for degradation in breast cancer. Consistently, using in vitro and in vivo models, we demonstrated that MLN4924, a specific small molecule inhibitor of NEDDylation, can restore ERRß expression and culminate in a reduction in cell proliferation and migration of breast cancer cells. We also showed that increased ERRß expression promotes the upregulation of its target genes, including the tumour suppressors p21Cip1/Waf1 and E-cadherin, involved in cell proliferation and migration arrest at the gene promoter level. Interestingly, this tumour suppressive role of ERRß does not depend on the expression of ERα in breast cancer. Moreover, our data revealed that the ERRß recruits the transcription co-activator p300 to its targeted gene promoters to upregulate their expression. Collectively, our work revealed that restoration of ERRß expression using the NEDDylation inhibitor MLN4924 can be a novel and effective strategy for breast cancer treatment.

Estrogen receptor α dependent regulation of estrogen related receptor ß and its role in cell cycle in breast cancer.

BACKGROUND: Breast cancer (BC) is highly heterogeneous with ~ 60-70% of estrogen receptor positive BC patient's response to anti-hormone therapy. Estrogen receptors (ERs) play an important role in breast cancer progression and treatment. Estrogen related receptors (ERRs) are a group of nuclear receptors which belong to orphan nuclear receptors, which have sequence homology with ERs and share target genes. Here, we investigated the possible role and clinicopathological importance of ERRß in breast cancer. METHODS: Estrogen related receptor ß (ERRß) expression was examined using tissue microarray slides (TMA) of Breast Carcinoma patients with adjacent normal by immunohistochemistry and in breast cancer cell lines. In order to investigate whether ERRß is a direct target of ERα, we investigated the expression of ERRß in short hairpin ribonucleic acid knockdown of ERα breast cancer cells by western blot, qRT-PCR and RT-PCR. We further confirmed the binding of ERα by electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP), Re-ChIP and luciferase assays. Fluorescence-activated cell sorting analysis (FACS) was performed to elucidate the role of ERRß in cell cycle regulation. A Kaplan-Meier Survival analysis of GEO dataset was performed to correlate the expression of ERRß with survival in breast cancer patients. RESULTS: Tissue microarray (TMA) analysis showed that ERRß is significantly down-regulated in breast carcinoma tissue samples compared to adjacent normal. ER + ve breast tumors and cell lines showed a significant expression of ERRß compared to ER-ve tumors and cell lines. Estrogen treatment significantly induced the expression of ERRß and it was ERα dependent. Mechanistic analyses indicate that ERα directly targets ERRß through estrogen response element and ERRß also mediates cell cycle regulation through p18, p21cip and cyclin D1 in breast cancer cells. Our results also showed the up-regulation of ERRß promoter activity in ectopically co-expressed ERα and ERRß breast cancer cell lines. Fluorescence-activated cell sorting analysis (FACS) showed increased G0/G1 phase cell population in ERRß overexpressed MCF7 cells. Furthermore, ERRß expression was inversely correlated with overall survival in breast cancer. Collectively our results suggest cell cycle and tumor suppressor role of ERRß in breast cancer cells which provide a potential avenue to target ERRß signaling pathway in breast cancer. CONCLUSION: Our results indicate that ERRß is a negative regulator of cell cycle and a possible tumor suppressor in breast cancer. ERRß could be therapeutic target for the treatment of breast cancer.