FAP-1 in pancreatic cancer cells: functional and mechanistic studies on its inhibitory role in CD95-mediated apoptosis.
In this study we investigated the functional role of FAP-1 as a potential inhibitor of CD95 (Fas, APO-1)-mediated apoptosis in pancreatic cancer cells. Stable transfection of the CD95-sensitive, FAP-1-negative cell line Capan-1 with an FAP-1 cDNA resulted in a strongly decreased sensitivity to CD95-induced apoptosis, as measured by DNA fragmentation and caspase-3 activity. Inhibition of cellular protein tyrosine phosphatases with orthovanadate dose-dependently increased CD95-induced apoptosis in CD95-resistant FAP-1-positive Panc89 and Capan-1-FAP-1 cells almost to the level seen in wild-type Capan-1 cells. Blocking the CD95/FAP-1 interaction in Panc89 cells by cytoplasmic microinjection of a synthetic tripeptide mimicking the C terminus of CD95 resulted in a mean 5.5-fold increase in apoptosis compared to cells that received a control peptide. Using confocal laser scanning microscopy we show that in Panc89 cells FAP-1 is mainly associated with the Golgi complex and with peripheral vesicles. FAP-1 displayed enhanced colocalization with CD95 upon CD95 stimulation in the Golgi complex but not in surface-associated vesicles. This correlated with a decrease in plasma membrane staining for CD95 as determined by FACS analysis. Inhibition of Golgi anterograde transport by brefeldin A abolished the anti-CD95-induced colocalization of FAP-1 and CD95 as well as the decrease in cell-surface-associated CD95. Finally, we demonstrate by immunohistochemistry that FAP-1 is strongly expressed in tumor cells from pancreatic carcinoma tissues. Taken together, these results show that FAP-1 can protect pancreatic carcinoma cells from CD95-mediated apoptosis, probably by preventing anti-CD95-induced translocation of CD95 from intracellular stores to the cell surface.
Ungefroren
H
H
Clinic for General Surgery and Thoracic Surgery, Christian-Albrechts-University, Kiel, Germany.
Kruse
M L
ML
Trauzold
A
A
Roeschmann
S
S
Roeder
C
C
Arlt
A
A
Henne-Bruns
D
D
Kalthoff
H
H
D000230:Adenocarcinoma; D017209:Apoptosis / Q000276:immunology*; D020126:Brefeldin A / Q000494:pharmacology; D002352:Carrier Proteins / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D022162:Cytoplasmic Vesicles / Q000737:chemistry / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic; D006056:Golgi Apparatus / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D019169:Jurkat Cells; D010190:Pancreatic Neoplasms; D010446:Peptide Fragments / Q000494:pharmacology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D054595:Protein Tyrosine Phosphatase, Non-Receptor Type 13; D017027:Protein Tyrosine Phosphatases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism*; D019014:fas Receptor / Q000032:analysis / Q000378:metabolism*
D002352:Carrier Proteins; D010446:Peptide Fragments; D011500:Protein Synthesis Inhibitors; D019014:fas Receptor; D020126:Brefeldin A; C516742:PTPN13 protein, human; D054595:Protein Tyrosine Phosphatase, Non-Receptor Type 13; D017027:Protein Tyrosine Phosphatases