Androgens activate mitogen-activated protein kinase via epidermal growth factor receptor/insulin-like growth factor 1 receptor in the mouse PC-1 cell line

Abstract

Androgens are the primary regulators of epididymal structure and functions. In the classical view of androgen action, binding of androgen to the intracellular androgen receptor (AR) produces the receptor-steroid complex that has high affinity for DNA response elements and regulates the transcription of target genes. In this study, we demonstrate that in epididymal cells, 5a-dihydrotestosterone (DHT) can cause an alternative and rapid response that is independent of AR-DNA interactions and is mediated by activation of signaling pathways through the AR. We examined changes in AKT and extracellular signal-regulated protein kinases (ERK1/2) activation at early time points after DHT supplementation in the mouse proximal caput epididymis-1 cell line. DHT had no significant effect on AKT activation at any time point. However, DHT activated the ERK pathway as early as at 1 min, the pathway remained activated at 10 min, but activation was not sustained at later time points. Interestingly, ERK activation was blocked by hydroxyflutamide (HF), indicating that early ERK activation was an AR-mediated response. DHT phosphorylates steroid receptor co-activator (SRC) kinase, and this activation was required for the ERK response. EGFR and IGF1R were downstream of SRC, and these two receptors together contributed to enhance ERK and cAMP response elementbinding protein (CREB) phosphorylation. We postulate that this rapid action of androgen may ultimately act to modulate the transcription of genes regulated by AR in the nucleus. These results support the hypothesis that DHT can activate a pathway involving the sequential activation of MEK, ERK1/2, and CREB through the EGFR/IGF1R in an epididymal cell line.

Estrogen receptor ESR1 mediates activation of ERK1/2, CREB, and ELK1 in the corpus of the epididymis

“…Overlapping and interconnected actions of E 2 (in the present study), androgens (Hamzeh & Robaire 2011), and growth factors (reviewed by Cotton et al 2008) on ERK1/2 activation may occur in the epididymis. An imbalance of these actions may result in infertility, and much remains to be learned about the complex interplay of hormonal actions in the epididymis.…”

“…Both steroids induce the interaction of ESR1 and androgen receptor (AR) with SRC as well as a conformational change that activates this kinase in breast cancer MCF-7 and prostate cancer LNCaP cells (Migliaccio et al 2000; reviewed by Migliaccio et al 2005). An SRC-dependent ERK activation by 5a-dihydrotestosterone (DHT) has also been described in the mouse proximal caput epididymis PC-1 cell line (Hamzeh & Robaire 2011). The phosphorylation of ERK1/2 induced by the activation of ESR1 was blocked by pretreatment with a non-receptor tyrosine kinase SRC inhibitor (PP2) in the corpus of the epididymis, which indicates that SRC is involved in this signal transduction.…”

“…Effects of the E 2 and ESR1-selective agonist PPT on the expression and phosphorylation of ERK1 and ERK2 in the corpus of the epididymis EGF receptor is present in the epididymis (Hamzeh & Robaire 2011). Therefore, EGF (10 ng/ml, 5 min) was used as a positive control for the phosphorylation of ERK1/2 in the corpus of the epididymis (Fig.…”

“…In fact, there is substantial evidence of cross-talk between the ERK1/2 and the PI3K/AKT pathways, either positively by cross-activation or negatively by crossinhibition, depending on the cellular environment (reviewed by Mendoza et al 2011). PI3K-mediated activation of the ERK1/2 pathway induced by relaxin in rat Sertoli cells (Nascimento et al 2012) and by DHT in the mouse proximal caput epididymis PC-1 cell line (Hamzeh & Robaire 2011) has been shown.…”

“…In preliminary experiments, we tested different concentrations of the inhibitors (PP2, AG1478, U0126, and Wortmannin), and the best concentrations were similar to those previously used in rat Sertoli cells (Lucas et al 2008, Royer et al 2012 and mouse proximal caput epididymis-1 cells (Hamzeh & Robaire 2011). All of the controls and treated tissues were processed in exactly the same way, and they remained in the nutrient solution for the same total incubation time.…”