Estrogen-related receptor gamma (ERRγ) is a critical regulator of mitochondrial metabolism and has been implicated in promoting resistance to tamoxifen in estrogen receptor-positive (ER+) breast cancer. Despite the clinical success of selective estrogen receptor modulators like tamoxifen, acquired resistance remains a significant challenge, affecting nearly half of treated patients. Unlike traditional resistance mechanisms involving loss or mutation of ER, tamoxifen resistance often arises through alterations in proliferative and survival signaling pathways. This study investigates how the ERK/MAPK kinase cascade regulates ERRγ expression and function, ultimately contributing to tamoxifen resistance.
We demonstrate that active ERK directly stabilizes ERRγ protein levels in ER+ breast cancer cells. Pharmacological inhibition of MEK with U0126 significantly reduces exogenous ERRγ expression, while inhibitors of JNK or p38 have no effect, indicating specificity for the ERK pathway. Furthermore, activation of MEK by a constitutively active mutant form (MEKDD) increases ERRγ levels, as does co-expression of wild-type ERK2 or stimulation with EGF. These findings confirm that ERK signaling positively regulates ERRγ abundance.
To identify functional phosphorylation sites, we analyzed the ERRγ sequence for potential ERK consensus motifs (S/T-P). Four serine residues met this criterion: S45, S57, S81, and S219. Using site-directed mutagenesis, we generated a triple-serine mutant (S57,81,219A). This mutant exhibited a 70% reduction in protein levels compared to wild-type ERRγ. Notably, MEKDD failed to enhance the mutant protein, and U0126 did not further reduce its levels, indicating that these three serines are essential for ERK-mediated stabilization of ERRγ.
Functional assays revealed that the S57,81,219A mutant is impaired in its ability to confer tamoxifen resistance. In MCF7 cells transfected with wild-type ERRγ, 4-hydroxytamoxifen (4HT) treatment induced only partial inhibition of DNA synthesis, whereas the mutant had no protective effect. Additionally, wild-type ERRγ suppressed the 4HT-induced upregulation of cyclin-dependent kinase inhibitors p21 and p27—key mediators of cell cycle arrest—but the mutant failed to do so.Recombinant Murine Interleukin-6Species Analysis of Rb phosphorylation showed that wild-type ERRγ promoted a shift toward hyperphosphorylated Rb, consistent with G1 progression, while the mutant did not alter Rb dynamics.GPA33 ProteinBiological Activity
Transcriptional activity assays confirmed that the S57,81,219A mutant exhibits reduced activity at canonical ER response elements (ERE), estrogen-related response elements (ERRE), and a hybrid ERRE/ERE site.PMID:34100241 The impairment was more pronounced in SUM44 cells than in MCF7 cells, suggesting context-dependent regulation likely influenced by differences in endogenous ER levels and co-regulatory proteins.
In summary, this study establishes ERK/MAPK as a key upstream regulator of ERRγ in ER+ breast cancer. Phosphorylation of S57, S81, and S219 by ERK stabilizes ERRγ protein and enhances its transcriptional activity, enabling it to promote cell cycle progression and bypass tamoxifen-induced growth inhibition. These findings reveal a novel mechanism linking oncogenic kinase signaling to metabolic reprogramming via an orphan nuclear receptor, offering new targets for overcoming tamoxifen resistance.MedChemExpress (MCE) offers a wide range of high-quality research chemicals and biochemicals (novel life-science reagents, reference compounds and natural compounds) for scientific use. We have professionally experienced and friendly staff to meet your needs. We are a competent and trustworthy partner for your research and scientific projects.Related websites: https://www.medchemexpress.com