pRb suppresses camptothecin-induced apoptosis in human osteosarcoma Saos-2 cells by inhibiting c-Jun N-terminal kinase.
This paper studies the cytotoxic effect induced by the topoisomerase I inhibitor camptothecin in human osteosarcoma Saos-2 cells, which lack p53 and contain a non-functional form of the product of the retinoblastoma gene, pRb. Cytotoxicity induced by camptothecin was dose- and time-dependent; the treatment with 100 nM camptothecin reduced cell viability by 50% at 32 h and by 75% at 72 h of exposure. The cytotoxic effect was caused by apoptosis, as ascertained by morphological evidence, acridine orange-ethidium bromide staining and flow cytometric analysis. Apoptosis was accompanied by both the activation of caspase-3 and the fragmentation of poly(ADP-ribose) polymerase. Treatment with camptothecin caused a threefold increase in the activity of c-Jun N-terminal kinase (JNK) and an eightfold increase in the level of phosphorylated c-Jun. The introduction of the RB gene into Saos-2 cells reduced the rate of cell growth. Moreover, stable clones of transfected cells were resistant to camptothecin. Exposure to 100 nM camptothecin for 72 h reduced the viability of transfected cells by only 10%; moreover, very modest effects were observed on the activity of JNK as well as on the level of phosphorylated c-Jun. The results reported in this paper support the conclusion that the expression of wild-type pRb in Saos-2 cells exerts an anti-apoptotic influence through the control of JNK activity.
Lauricella
M
M
Department of Cell and Developmental Biology, Section of Biochemistry, University of Palermo, Policlinico, Via del Vespro 129, 90127, Palermo, Italy.
Calvaruso
G
G
Carabillò
M
M
D'Anneo
A
A
Giuliano
M
M
Emanuele
S
S
Vento
R
R
Tesoriere
G
G
D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002166:Camptothecin / Q000037:antagonists & inhibitors* / Q000633:toxicity; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D048429:Cell Size / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D004264:DNA Topoisomerases, Type I / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D005434:Flow Cytometry; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D018384:Oxidative Stress / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D016160:Retinoblastoma Protein / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D059004:Topoisomerase I Inhibitors; D014162:Transfection; D014407:Tumor Cells, Cultured
D016755:Proto-Oncogene Proteins c-jun; D016160:Retinoblastoma Protein; D059004:Topoisomerase I Inhibitors; D006861:Hydrogen Peroxide; D011065:Poly(ADP-ribose) Polymerases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D004264:DNA Topoisomerases, Type I; D005978:Glutathione; D002166:Camptothecin