Participation of various kinases in staurosporine induced apoptosis of RAW 264.7 cells.
Staurosporine induced apoptosis of RAW 264.7 cells, a mouse macrophage-like cell line, as determined by DNA fragmentation, the increase of annexin V-stained cells, and the cleavage of poly(ADP-ribose)polymerase (PARP), a substrate of caspase. Analysis of the increase in the percentage of sub-G(1) cells revealed that the DNA fragmentation occurred in a time- and concentration-dependent manner at 0.021-2.1 microM of staurosporine. Staurosporine induced phosphorylation of p38 mitogen-activated protein kinase (MAPK) but suppressed spontaneous phosphorylation of p44/42 MAPK. The p38 MAPK inhibitor SB203580, the MAPK/extracellular signal-regulated kinase kinase inhibitor PD98059 and the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 potentiated the staurosporine-induced PARP cleavage and DNA fragmentation. The protein kinase A (PKA) inhibitor H-89 potentiated the staurosporine-induced DNA fragmentation without potentiating the PARP cleavage. In contrast, the protein kinase C (PKC) inhibitor Ro-31-8425 suppressed the PARP cleavage and DNA fragmentation. These findings suggested that staurosporine induces apoptosis via the caspase cascade in RAW 264.7 cells. The staurosporine-induced apoptosis is positively regulated by PKC, negatively regulated by p38 MAPK, p44/42 MAPK and PI3K via the caspase cascade, and negatively regulated by PKA without regulation of caspase activation.
Yamaki
Kouya
K
Laboratory of Pathophysiological Biochemistry, Graduate School of Pharmaceutical Sciences, Tohoku University, Aoba Aramaki, Aobaku, Sendai, Miyagi 980-8578, Japan.
Hong
Jangja
J
Hiraizumi
Kenji
K
Ahn
Jong Woon
JW
Zee
OkPyo
O
Ohuchi
Kazuo
K
D000818:Animals; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D053938:DNA Fragmentation / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005419:Flavonoids / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009025:Morpholines / Q000494:pharmacology; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D019311:Staurosporine / Q000494:pharmacology*; D013449:Sulfonamides; D048051:p38 Mitogen-Activated Protein Kinases
D002867:Chromones; D004791:Enzyme Inhibitors; D005419:Flavonoids; D007546:Isoquinolines; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D013449:Sulfonamides; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D011493:Protein Kinase C; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D020169:Caspases; D019311:Staurosporine; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one