Fas-associated phosphatase-1 promotes Fas-mediated apoptosis in human colon cancer cells: novel function of FAP-1.
OBJECTIVE
Fas-associated phosphatase-1 (FAP-1) has been thought as an inhibitor in Fas-mediated apoptosis. Here, we investigated the role of FAP-1 in Fas-mediated apoptosis of human colon cancer cells.


METHODS
The viability of four colon cancer cell lines treated with agonistic anti-Fas antibody was determined using WST-1 assay and cell death detection ELISA. pRc/CMV-FAP-1 was transfected to a FAP-1-negative, Fas-resistant colon cancer cell line SW480 by lipofection and the clones expressing FAP-1 protein were selected by limiting dilution. In the clones, expression of 550 genes was analyzed by cDNA microarrays. Protein expression of FAP-1 and molecules related to apoptosis was examined by western blot.


RESULTS
We obtained two FAP-1 overexpressed clones which were much more susceptible to Fas-mediated apoptosis than control cells. In the clones, caspase 8 and caspase 3 were fully activated by agonistic anti-Fas antibody treatment. Bcl-2 family proteins were not related to the high susceptibility of these clones, because caspase 9 was not activated. Transfection of FAP-1 did not suppress the survival actions of insulin-like growth factor (IGF-1) which enhanced survival signal through Akt phosphorylation. Upregulation in 21 genes and downregulation in 29 genes was revealed by cDNA arrays. We confirmed protein expression of p21 and phosphorylated p21 were much more enhanced in the clones than in control cells.


CONCLUSIONS
Overexpression of FAP-1 enhanced susceptibility to Fas-mediated apoptosis in SW480 and upregulation of p21 may contribute to this phenomenon. Our results indicate a novel function of FAP-1 in Fas-mediated apoptosis of human colon cancer cells.
Miyazaki
Takako
T
Third Department of Internal Medicine, Division of Transfusion Medicine and Cell Therapy, Toyama Medical and Pharmaceutical University, Toyama, Japan.
Atarashi
Yoshinari
Y
Yasumura
Satoshi
S
Minatoya
Isao
I
Ogawa
Kohei
K
Iwamoto
Masaya
M
Minemura
Masami
M
Shimizu
Yukihiro
Y
Sato
Taka-Aki
TA
Watanabe
Akiharu
A
Sugiyama
Toshiro
T
D000906:Antibodies; D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins / Q000235:genetics / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival; D003110:Colonic Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000235:genetics / Q000378:metabolism; D020869:Gene Expression Profiling; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010766:Phosphorylation / Q000187:drug effects; D054645:Protein Phosphatase 1; D054595:Protein Tyrosine Phosphatase, Non-Receptor Type 13; D017027:Protein Tyrosine Phosphatases / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection; D019014:fas Receptor / Q000276:immunology / Q000378:metabolism*
D000906:Antibodies; D051017:Apoptosis Regulatory Proteins; C493956:CDKN1A protein, human; C481004:CH-11 anti-fas antibody, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; C491901:FAS protein, human; C112037:FCMR protein, human; D008565:Membrane Proteins; D012333:RNA, Messenger; D019014:fas Receptor; D007334:Insulin-Like Growth Factor I; D051057:Proto-Oncogene Proteins c-akt; D054645:Protein Phosphatase 1; C516742:PTPN13 protein, human; D054595:Protein Tyrosine Phosphatase, Non-Receptor Type 13; D017027:Protein Tyrosine Phosphatases; D053148:Caspase 3; D053181:Caspase 8