Soluble factors derived from stroma activated androgen receptor phosphorylation in human prostate LNCaP cells: roles of ERK/MAP kinase.
BACKGROUND
Accumulated evidence suggests stromal-epithelial interactions are critical to the progression of prostate cancer. In this study, we characterized AR phosphorylation in LNCaP cells co-cultured with the conditioned medium (CM) from human prostate stromal fibroblasts.


METHODS
CM harvested from prostate stromal fibroblasts was added to LNCaP cells, and both anchorage-dependent and -independent growth was determined. Status of AR phosphorylation at Ser-81 and Ser-213 was assessed by immunoblot analysis. ERK kinase activity was measured using MBP-2 protein as the substrate.


RESULTS
The growth of LNCaP cells on plastic dishes increased by 1.7-fold upon exposure to stromal CM or androgen, and their combination resulted in additive growth (2.4-fold). Anchorage-independent growth of LNCaP cells in soft agar, however, was induced synergistically at 80-fold by both stromal CM and androgen. Stromal CM or androgen alone induced LNCaP cell growth by 10- and 26-fold, respectively. We observed ERK kinase inhibitor, U0126, but not phosphatidylinositol 3-kinase (PI-3K), LY294002, or protein kinase A (PKA) inhibitor, H-89, inhibited stromal CM or androgen-induced PSA promoter luciferase activities, and anchorage-independent growth of LNCaP cells. Our results demonstrated for the first time how stromal CM acts in synergy with androgen by activation of ERK kinase and AR phosphorylation at Ser-81 but not Ser-213, for AR-regulated PSA promoter and anchorage-independent growth of human prostate cancer cells.


CONCLUSIONS
A stromal factor-activated ERK pathway mediated by AR phosphorylation at Ser-81 could be responsible for stimulating the growth of human prostate cancer cells.
Shigemura
Katsumi
K
Molecular Urology and Therapeutics Program, Department of Urology, Winship Cancer Institute, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Isotani
Shuji
S
Wang
Ruoxiang
R
Fujisawa
Masato
M
Gotoh
Akinobu
A
Marshall
Fray F
FF
Zhau
Haiyen E
HE
Chung
Leland W K
LW
D000728:Androgens / Q000378:metabolism; D002450:Cell Communication / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology*; D008297:Male; D010766:Phosphorylation / Q000502:physiology; D017430:Prostate-Specific Antigen / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000378:metabolism* / Q000473:pathology*; D011944:Receptors, Androgen / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D012995:Solubility; D013234:Stem Cells; D017154:Stromal Cells / Q000166:cytology / Q000378:metabolism*; D014407:Tumor Cells, Cultured
C410798:AR protein, human; D000728:Androgens; D017077:Culture Media, Conditioned; D011944:Receptors, Androgen; D012694:Serine; D008156:Luciferases; D048049:Extracellular Signal-Regulated MAP Kinases; D017430:Prostate-Specific Antigen