ERα-36 regulates progesterone receptor activity in breast cancer

Abstract

Background: Alterations in estrogen and progesterone signaling, via their respective receptors, estrogen receptor alpha (ERα) and progesterone receptor (PR), respectively, are largely involved in the development of breast cancer (BC). The recent identification of ERα-36, a splice variant of ERα, has uncovered a new facet of this pathology. Although ERα-36 expression is associated with poor prognosis, metastasis development, and resistance to treatment, its predictive value has so far not been associated with a BC subtype and its mechanisms of action remain understudied. Methods: To study ERα-36 expression in BC specimens, we performed immunochemical experiments. Next, the role of ERα-36 in progesterone signaling was investigated by generating KO clones using the CRISPR/CAS9 technology. PR signaling was also assessed by proximity ligation assay, Western blotting, RT-QPCR, and ChIP experiments. Finally, proliferation assays were performed with the IncuCyte technology and migration experiments using scratch assays. Results: Here, we demonstrate that ERα-36 expression at the plasma membrane is correlated with a reduced diseasefree survival in a cohort of 160 BC patients, particularly in PR-positive tumors, suggesting a crosstalk between ERα-36 and PR. Indeed, we show that ERα-36 interacts constitutively with PR in the nucleus of tumor cells. Moreover, it regulates PR expression and phosphorylation on key residues, impacting the biological effects of progesterone. Conclusions: ERα-36 is thus a regulator of PR signaling, interfering with its transcriptional activity and progesteroneinduced anti-proliferative effects as well as migratory capacity. Hence, ERα-36 may constitute a new prognostic marker as well as a potential target in PR-positive BC.

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“…However, in another Chinese patient cohort with ER-negative breast cancer, ERα36 was not found to have any correlation with clinicopathological characteristics, and its inhibition in breast cancer cell lines was reported to increase their sensitivity to paclitaxel [ 51 ]; however, membrane/submembrane expression was not examined separately in this study. A recent study also reports the direct interaction of ERα36 with PR, with the first one inducing the expression of the latter and thus affecting its activity in a manner that increases the aggressiveness of breast cancer cells and leading to poorer patient prognosis [ 13 ]. However, in the last study, all PR-positive patients were also ERα66-positive, complicating the interpretation of these findings, while an unexpected better prognosis of PR-negative patients expressing high levels of ERα36 was observed.…”

“…Deriving from an alternative transcriptional initiation at the first intron, it contains exons 2–6 of the classic ERα and a unique 27-amino acid C′-terminal sequence, thus missing transcriptional activation domain AF1 and part of AF2, but retaining the DNA-binding domain, the dimerization capacity, and most of the sequence of ERα66 critical for ligand binding [ 6 ]. We have previously reported that ERα36 is expressed in breast cancer cell lines and in the cancer tissues of a cohort with triple-negative breast cancer (TNBC) patients, where its membrane localization is a good prognostic indicator [ 11 ], although controversial results regarding the clinical significance of this isoform exist [ 12 , 13 , 14 ].…”

The Role of ERα36 in Development and Tumor Malignancy

“…A recent study confirmed that the expression of ERα36 was of poor prognosis in PR-positive tumors. However, this expression was significantly associated with longer survival in PR-negative tumors, suggesting a crosstalk between ERα36 and PR in breast cancer [55]. Therefore, ERα36 could become a new prognostic marker to discriminate a subset of PR tumors, generally associated with a good prognosis that may develop metastases.…”

“…However, so far, ERα36 has been shown to regulate only ERα activity [11]. Recently, Konan and colleagues [55], showed that ERα36 binds to progesterone receptor (PR) and interferes with progesterone signaling. Indeed, ERα36 modulates PR expression, phosphorylation, and transcriptional activity.…”

“…Interestingly, ERα36 is involved in the action of progesterone on cell proliferation and migration. Moreover, ERα36 expression is associated with a reduced metastasis disease-free survival in PR-positive tumors [55].…”