Androgen Receptor Mediates Non-genomic Activation of Phosphatidylinositol 3-OH Kinase in Androgen-sensitive Epithelial Cells

Abstract

Androgens are known to modulate many cellular processes such as cell growth and survival by binding to the androgen receptor (AR) and activating the transcription of target genes. Recent data suggested that AR can also mediate non-transcriptional actions outside the nucleus in addition to its ligand-inducible transcription factor function. Here, we describe a transcription-independent activation of the phosphatidylinositol 3-OH kinase (PI3-K) signaling pathway by androgens. Using non-transformed androgen-sensitive epithelial cells, we show that androgens enhance the PI3-K activity by promoting accumulation of phosphoinositide-3-P phospholipids in vitro. This activation is found in conjunction with an increased time-dependent phosphorylation of the downstream kinase AKT/protein kinase B on both Ser 473 and Thr 308 residues. Hormone-stimulated phosphorylation of AKT requires AR since incubation with the anti-androgen bicalutamide completely abolishes the androgen-stimulated AKT phosphorylation. Accordingly, we show that androgens increase AKT phosphorylation level in prostatic carcinoma PC3 cells only once they have been transfected with AR. Downstream, androgens enhance phosphorylation of transcription factor FKHR (Forkhead in rhabdomyosarcoma)-L1 and proapoptotic Bad protein and promote cell survival as they can counteract an apoptotic process. We also report that non-genomic effects of androgens are based on direct interaction between AR and the p85␣ regulatory subunit of class I(A) PI3-K. Together, these novel findings point out an important and physiologically relevant link between androgens and the PI3-K/AKT signaling pathway in governing cell survival.

Upregulation of VEGF-C by androgen depletion: the involvement of IGF-IR-FOXO pathway

“…A recent report suggested that androgen stimulation of LNCaP cells cultured in media containing charcoal-dextran-treated serum leads to an increase in IGF-IR protein expression (Arnold et al, 2005). The activation of PI3K pathway by androgen was also reported previously (Baron et al, 2004;Castoria et al, 2004;Kang et al, 2004). These reports, together with our observation of higher VEGF-C mRNA levels after inhibiting IGF-IR signaling supports the conclusion that androgen deprivation leads to a decrease in IGF-IR expression that in turn increases the transcription of VEGF-C.…”

“…SIRT-1DN protein overexpression was detected by Western blot with the anti-FLAG antibody (Figure 4e). Since androgen withdrawal leads to inhibition of PI3K-AKT pathway (Baron et al, 2004;Castoria et al, 2004;Kang et al, 2004) and therefore potentially activates FOXO-1, we tested whether dominant-negative form of SIRT-1 could abrogate the increase in VEGF-C expression in LNCaP due to androgen withdrawal. Our result (Figure 4f) clearly indicated the involvement of SIRT-1 in androgenregulated VEGF-C transcription.…”

Androgen Action During Prostate Carcinogenesis

“…In addition to the classic genomic effects of sex steroids, accumulating data have also shown the importance of nongenomic effects (4)(5)(6)(7). For instance, androgen treatment results in the association of AR with Src kinase, and thereby activates Src/Raf-1/Erk pathway, leading to cell proliferation and survival (8).…”

“…Inhibition of AKT with a dominant-negative AKT or a PI3K inhibitor significantly attenuates androgen-induced cell proliferation (4,5). Androgen-induced AKT activation requires the AR, but deletion of the ligand-binding domain of AR does not abolish it, indicating that this regulation is independent of AR transcriptional activity.…”

“…Androgen-induced AKT activation requires the AR, but deletion of the ligand-binding domain of AR does not abolish it, indicating that this regulation is independent of AR transcriptional activity. Indeed, androgen-bound AR physically binds to the p85α regulatory subunit of PI3K and thereby activates PI3K/AKT (4,5). This is truly a two-way crosstalk, since AR expression and activity are also regulated by PI3K/AKT (67)(68)(69).…”