Neutral endopeptidase inhibits prostate cancer cell migration by blocking focal adhesion kinase signaling.
Neutral endopeptidase 24.11 (NEP, CD10) is a cell-surface enzyme expressed by prostatic epithelial cells that cleaves and inactivates neuropeptides implicated in the growth of androgen-independent prostate cancer (PC). NEP substrates such as bombesin and endothelin-1 induce cell migration. We investigated the mechanisms of NEP regulation of cell migration in PC cells, including regulation of phosphorylation on tyrosine of focal adhesion kinase (FAK). Western analyses and cell migration assays revealed an inverse correlation between NEP expression and the levels of FAK phosphorylation and cell migration in PC cell lines. Constitutively expressed NEP, recombinant NEP, and induced NEP expression using a tetracycline-repressive expression system inhibited bombesin- and endothelin-1-stimulated FAK phosphorylation and cell migration. This results from NEP-induced inhibition of neuropeptide-stimulated association of FAK with cSrc protein. Expression of a mutated catalytically inactive NEP protein also resulted in partial inhibition of FAK phosphorylation and cell migration. Coimmunoprecipitation experiments show that NEP associates with tyrosine-phosphorylated Lyn kinase, which then binds the p85 subunit of phosphatidylinositol 3-kinase (PI3-K) resulting in an NEP-Lyn-PI3-K protein complex. This complex competitively blocks FAK-PI3-K interaction, suggesting that NEP protein inhibits cell migration via a protein-protein interaction independent of its catalytic function. These experiments demonstrate that NEP can inhibit FAK phosphorylation on tyrosine and PC cell migration through multiple pathways and suggest that cell migration which contributes to invasion and metastases in PC cells can be regulated by NEP.
Sumitomo
M
M
Urologic Oncology Research Laboratory, Department of Urology, and. Department of Pathology, Weill Medical College of Cornell University, New York, New York, USA.
Shen
R
R
Walburg
M
M
Dai
J
J
Geng
Y
Y
Navarro
D
D
Boileau
G
G
Papandreou
C N
CN
Giancotti
F G
FG
Knudsen
B
B
Nanus
D M
DM
D000818:Animals; D001839:Bombesin / Q000494:pharmacology; D019556:COS Cells; D002465:Cell Movement / Q000187:drug effects; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004274:DNA, Recombinant / Q000235:genetics / Q000378:metabolism; D019332:Endothelin-1 / Q000494:pharmacology; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D006801:Humans; D008297:Male; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D015260:Neprilysin / Q000235:genetics / Q000378:metabolism*; D063065:Organophosphonates / Q000494:pharmacology; D010649:Phenylalanine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D019061:src-Family Kinases / Q000378:metabolism
C086164:CGS 24592; D016895:Culture Media, Serum-Free; D004274:DNA, Recombinant; D019332:Endothelin-1; D008565:Membrane Proteins; D063065:Organophosphonates; D010649:Phenylalanine; D019869:Phosphatidylinositol 3-Kinases; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; D016392:Proto-Oncogene Proteins pp60(c-src); C060902:lyn protein-tyrosine kinase; D019061:src-Family Kinases; D015260:Neprilysin; D001839:Bombesin