Protein kinase Czeta mediated Raf-1/extracellular-regulated kinase activation by daunorubicin.
In light of the emerging concept of a protective function of the mitogen-activated protein kinase (MAPK) pathway under stress conditions, we investigated the influence of the anthracycline daunorubicin (DNR) on MAPK signaling and its possible contribution to DNR-induced cytotoxicity. We show that DNR increased phosphorylation of extracellular-regulated kinases (ERKs) and stimulated activities of both Raf-1 and extracellular-regulated kinase 1 (ERK1) within 10 to 30 minutes in U937 cells. ERK1 stimulation was completely blocked by either the mitogen-induced extracellular kinase (MEK) inhibitor PD98059 or the Raf-1 inhibitor 8-bromo-cAMP (cyclic adenosine monophosphate). However, only partial inhibition of Raf-1 and ERK1 stimulation was observed with the antioxidant N-acetylcysteine (N-Ac). Moreover, the xanthogenate compound D609 that inhibits DNR-induced phosphatidylcholine (PC) hydrolysis and subsequent diacylglycerol (DAG) production, as well as wortmannin that blocks phosphoinositide-3 kinase (PI3K) stimulation, only partially inhibited Raf-1 and ERK1 stimulation. We also observed that DNR stimulated protein kinase C zeta (PKCzeta), an atypical PKC isoform, and that both D609 and wortmannin significantly inhibited DNR-triggered PKCzeta activation. Finally, we found that the expression of PKCzeta kinase-defective mutant resulted in the abrogation of DNR-induced ERK phosphorylation. Altogether, these results demonstrate that DNR activates the classical Raf-1/MEK/ERK pathway and that Raf-1 activation is mediated through complex signaling pathways that involve at least 2 contributors: PC-derived DAG and PI3K products that converge toward PKCzeta. Moreover, we show that both Raf-1 and MEK inhibitors, as well as PKCzeta inhibition, sensitized cells to DNR-induced cytotoxicity.
Mas
Véronique Mansat-De
VM
Institut National de la Santé et de la Recherche Médicale U563, Institut Claudius Régaud, Toulouse, France. demas.v@chu-toulouse.fr
Hernandez
Hélène
H
Plo
Isabelle
I
Bezombes
Christine
C
Maestre
Nicolas
N
Quillet-Mary
Anne
A
Filomenko
Rodolphe
R
Demur
Cécile
C
Jaffrézou
Jean-Pierre
JP
Laurent
Guy
G
D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000111:Acetylcysteine / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology; D000903:Antibiotics, Antineoplastic / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D001952:Bridged-Ring Compounds / Q000494:pharmacology; D016923:Cell Death / Q000187:drug effects; D003630:Daunorubicin / Q000494:pharmacology*; D004075:Diglycerides / Q000096:biosynthesis; D004789:Enzyme Activation / Q000187:drug effects; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D007938:Leukemia; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009636:Norbornanes; D010713:Phosphatidylcholines / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf / Q000037:antagonists & inhibitors / Q000378:metabolism*; D017382:Reactive Oxygen Species / Q000494:pharmacology; D013859:Thiocarbamates; D013871:Thiones / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured; D000077191:Wortmannin; D000099221:Protein Kinase C zeta
D015124:8-Bromo Cyclic Adenosine Monophosphate; D000111:Acetylcysteine; D000730:Androstadienes; D000903:Antibiotics, Antineoplastic; D000975:Antioxidants; D001952:Bridged-Ring Compounds; D003630:Daunorubicin; D004075:Diglycerides; D005419:Flavonoids; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D009636:Norbornanes; D010713:Phosphatidylcholines; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D019908:Proto-Oncogene Proteins c-raf; D017382:Reactive Oxygen Species; D013859:Thiocarbamates; D013871:Thiones; D000077191:Wortmannin; D000099221:Protein Kinase C zeta; C046498:tricyclodecane-9-yl-xanthogenate; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one