Fas-induced apoptosis in human malignant melanoma cell lines is associated with the activation of the p34(cdc2)-related PITSLRE protein kinases.
The Cdc2L locus encoding the PITSLRE protein kinases maps to chromosome band 1p36 and consists of two duplicated and tandemly linked genes. The purpose of the present study was to determine whether diminution of PITSLRE kinases leads to deregulation of apoptosis. The human melanoma cell lines A375 (Cdc2L wild-type alleles) and UACC 1227 (mutant Cdc2L alleles) were tested with agonist anti-Fas monoclonal antibody. We found that exposure of these cells to anti-Fas for 24, 48, or 72 h resulted in differential sensitivity to Fas-induced apoptosis. In A375, cell death started at 24-48 h post-treatment, and it was maximal by 72 h. Conversely, UACC 1227 cells were resistant to Fas-mediated apoptosis. Induction of PITSLRE histone H1 kinase activity was observed in A375 anti-Fas treated but not in UACC 1227 cells. Also, the PITSLRE protein kinase activity in A375 anti-Fas-treated cells preceded maximal levels of apoptosis. Finally, fluorescence confocal microscopy revealed a nuclear localization of PITSLRE proteins in normal melanocytes and A375 cells but a cytoplasmic localization in UACC 1227 cells. The differences in PITSLRE protein and cellular localization between A375 and UACC 1227 cells appear to account for the differences in sensitivity of the two cells lines to anti-Fas and staurosporine. These observations suggest that alterations in PITSLRE gene expression and protein localization may result in the loss of apoptotic signaling.
Ariza
M E
ME
Arizona Cancer Center, Tucson, Arizona 85724, USA.
Broome-Powell
M
M
Lahti
J M
JM
Kidd
V J
VJ
Nelson
M A
MA
D000911:Antibodies, Monoclonal / Q000276:immunology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D016203:CDC2 Protein Kinase / Q000378:metabolism*; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D061945:Caspase Inhibitors; D002467:Cell Nucleus / Q000378:metabolism; D018844:Cyclin-Dependent Kinases; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation; D053222:Fas Ligand Protein; D006801:Humans; D008545:Melanoma / Q000201:enzymology / Q000378:metabolism / Q000473:pathology*; D008562:Membrane Glycoproteins / Q000378:metabolism; D008845:Microinjections; D011494:Protein Kinases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases; D019311:Staurosporine / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000276:immunology / Q000502:physiology*
D000911:Antibodies, Monoclonal; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D019014:fas Receptor; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase; C447612:CDK11a protein, human; D018844:Cyclin-Dependent Kinases; C505407:CASP3 protein, human; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D019311:Staurosporine