FAP-1 in pancreatic cancer cells: functional and mechanistic studies on its inhibitory role in CD95-mediated apoptosis
2001
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.
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