Stable Inhibition of Specific Estrogen Receptor α (ERα) Phosphorylation Confers Increased Growth, Migration/Invasion, and Disruption of Estradiol Signaling in MCF-7 Breast Cancer Cells

Abstract

Elevated phosphorylation of estrogen receptor α (ERα) at serines 118 (S118) and 167 (S167) is associated with favorable outcome for tamoxifen adjuvant therapy and may serve as surrogate markers for a functional ERα signaling pathway in breast cancer. It is possible that loss of phosphorylation at S118 and/or S167 could disrupt ERα signaling, resulting in aggressive ERα-independent breast cancer cells. To this end, MCF-7 breast cancer cells were stably transfected with an ERα-specific short hairpin RNA that reduced endogenous ERα. The resulting cell line was stably transfected with wild-type ERα (ER-AB cells), or ERα containing serine to alanine mutation at S118 or S167 (S118A cells and S167A cells, respectively). These stable cell lines expressed approximately equivalent ERα compared with parental MCF-7 cells and were evaluated for growth, morphology, migration/invasion, and ERα-regulated gene expression. S118A cells and S167A cells exhibited increased growth and migration/invasion in vitro. Forward- and side-scatter flow cytometry revealed that S167A cells were smaller in size, and both S118A and S167A cells exhibited less cellular complexity. S118A and S167A cells expressed pancytokeratin and membrane localization of β-catenin and did not express vimentin, indicating retention of epithelial lineage markers. Expression of ERα-target genes and other genes regulated by ERα signaling or involved in breast cancer were markedly altered in both S118A and S167A cells. In summary, attenuated phosphorylation of ERα at S118 and S167 significantly affected cellular physiology and behavior in MCF-7 breast cancer cells, resulting in increased growth, migration/invasion, compromised expression of ERα target genes, and markedly altered gene expression patterns.

Estrogen receptor α mediates proliferation of breast cancer MCF–7 cells via a p21/PCNA/E2F1‐dependent pathway

“…Huderson et al . reports that phosphorylation of ERα increased growth and migration/invasion in MCF‐7 breast cancer cells. Sisci et al .…”

“…However, the effects of ERa on migration and invasion of MCF-7 cells appear to be contradictory. Huderson et al [28] reports that phosphorylation of ERa increased growth and migration/invasion in MCF-7 breast cancer cells. Sisci et al [29] reported that cell adhesion on fibronectin and type IV collagen induced ERa-mediated transcription and reduced cell migration in MCF-7 and MDA-MB-231 cell lines expressing ERa.…”

Neddylation Inactivation Facilitates FOXO3a Nuclear Export to Suppress Estrogen Receptor Transcription and Improve Fulvestrant Sensitivity

“…It is reported that ER is transcriptionally activated by phosphorylation, which contributes to the regulation of multiple biological processes including hormone sensitivity, nuclear localization, DNA binding, protein/chromatin interactions, protein stability, and gene transcription (32,33). The most common phosphorylation sites reported by numerous laboratories are serine 118 (Ser118) and serine 167 (Ser167; ref.…”

A Ternary Mixture of Common Chemicals Perturbs Benign Human Breast Epithelial Cells More Than the Same Chemicals Do Individually

“…ERα phosphorylated at serine 118 (S118) in the Activation function-1 domain localizes preferentially at promoters of ER responsive genes ( Weitsman et al , 2006 ). This binding is critical for increased cell growth and resistance to apoptotic cell death ( Huderson et al , 2012 ), supporting the hypothesis that activation at this key phosphorylation site is a potential surrogate for a functional ERα signaling pathway in breast cancer. (4) Apoptosis evasion : Well recognized as a characteristic of cancer cells, induction of apoptotic cell death is a common objective of all forms of cancer treatment, i.e.…”