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17β-Estradiol activates GPER- and ESR1-dependent pathways inducing apoptosis in GC-2 cells, a mouse
Adele Chimento1, Rosa Sirianni, Ivan Casaburi
1Department of Pharmaco-Biology, University of Calabria, 87036 Arcavacata di Rende, Cosenza, Italy.
Abstract:
In mammals, spontaneous apoptosis is observed particularly in differentiating spermatogonia and in spermatocytes. 17β-Estradiol (E2) in primary rat pachytene spermatocytes (PS) binds estrogen receptor α (ESR1) and GPER to activate EGFR/ERK/c-Jun pathway leading to up regulation of proapoptotic factor bax. Aim of this study was to clarify the effector pathway(s) controlling spermatocytes apoptosis using as model GC-2 cells, an immortalized mouse pachytene spermatocyte-derived cell line, which reproduces primary cells responses to E2. In fact, in GC-2 cells we observed that ESR1 and GPER activation caused rapid ERK and c-Jun phosphorylation, bax up-regulation, events associated with apoptosis. We further investigated the apoptotic mechanism demonstrating that E2, as well as ESR1 and GPER specific agonists, induced sustained ERK, c-Jun and p38 phosphorylation, Cytochrome c release, caspase 3 and endogenous substrate Poly (ADP-ribose) polymerase (PARP) activation and increased expression of cell cycle inhibitor p21. When ESR1 or GPER expression was silenced, E2 was still able to decrease cell proliferation, only the concomitant silencing abolished E2 effect. These results indicate that GC-2 cells are a valid cell model to study E2-dependent apoptosis in spermatocytes and show that E2, activating both ESR1 and GPER, is able to induce an ERK1/2, c-Jun and p38-dependent mitochondrion apoptotic pathway in this cell type.
Insights
17β-Estradiol (E2) induces apoptosis in spermatocytes by activating estrogen receptor α (ESR1) and GPER. This triggers a mitochondrial pathway involving ERK1/2, c-Jun, and p38 phosphorylation, leading to cell death.
Area of Science:
- Reproductive biology
- Cell biology
- Endocrinology
Background:
- Spontaneous apoptosis occurs in mammalian spermatogonia and spermatocytes.
- 17β-Estradiol (E2) activates estrogen receptor α (ESR1) and GPER in primary rat pachytene spermatocytes.
- This activation initiates the EGFR/ERK/c-Jun pathway, upregulating the proapoptotic factor BAX.
Purpose of the Study:
- To elucidate the effector pathways controlling spermatocyte apoptosis.
- To validate GC-2 cells as a model for studying E2-dependent apoptosis in spermatocytes.
- To investigate the specific roles of ESR1 and GPER in E2-induced apoptosis.
Main Methods:
- Utilized GC-2 cells, an immortalized mouse pachytene spermatocyte cell line.
- Examined the effects of E2, ESR1, and GPER agonists on cellular signaling pathways.
- Assessed apoptosis markers including phosphorylation, cytochrome c release, caspase activation, and PARP cleavage.
- Employed gene silencing techniques to investigate receptor function.
Main Results:
- E2, ESR1, and GPER activation in GC-2 cells induced rapid ERK and c-Jun phosphorylation and BAX upregulation.
- E2 treatment led to sustained ERK, c-Jun, and p38 phosphorylation, cytochrome c release, caspase 3 and PARP activation, and increased p21 expression.
- Concomitant silencing of ESR1 and GPER abolished E2's effect on cell proliferation, indicating a dual receptor role.
Conclusions:
- GC-2 cells serve as a valid model for studying E2-dependent spermatocyte apoptosis.
- E2 induces apoptosis via a mitochondrial pathway dependent on ERK1/2, c-Jun, and p38 phosphorylation.
- Both ESR1 and GPER are crucial for E2-mediated apoptosis and proliferation inhibition in spermatocytes.
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