Abstract
We previously demonstrated that ectopic expression of neurotrophic peptide (NP) derived from saposin C promotes androgen receptor (AR) expression and transactivation in human prostate cancer cells. This prompted us to investigate how NP or saposin C can function in cells. We constructed plasmids expressing saposin C or a chimeric peptide of a viral TAT transduction domain and saposin C (TAT‐saposin C) with His‐tag. Intracellular localization of saposin C and NP was predominantly shown in transfected cells, while TAT‐saposin C was detected around membrane and in cytosol by immunofluorescence staining. Furthermore, induction of the AR expression and activation of the AR transcriptional function were observed in cells transfected with saposin C or TAT‐saposin C, compared to control cells transfected with an empty plasmid. The effects of saposin C and TAT‐saposin C on AR activity were examined in the presence of inhibitors of GPCR, MAPK1/2, and PI3K/Akt. Interestingly, we found that these inhibitors only affect AR activities in cells with TAT‐saposin C expression but not with saposin C expression. Immunostaining images showed that co‐localization of saposin C, Src, and the AR occurred in transfected cells. Physical interactions of saposin C/NP, Src, and the AR were then demonstrated by co‐immunoprecipitation assays. Blockage of Src activity by specific inhibitor led to a decrease in the saposin C‐mediated enhancement of AR transactivity, suggesting that intracellular expression of saposin C caused stimulation of AR expression and activity by associations with Src in LNCaP cells. This effect may not be mediated by GPCR. J. Cell. Biochem. 112: 818–828, 2011. © 2010 Wiley‐Liss, Inc.