Multiple signal transduction pathways mediate c-Jun protein phosphorylation.
A variety of protein kinases, including pp42 and pp54 mitogen-activated protein (MAP) kinases, p34cdc2, and a partially purified protein kinase from 4 beta-phorbol 12-myristate 13 alpha-acetate (PMA)-treated U937 cells have been shown to phosphorylate the NH2-terminal activation domain of c-Jun in vitro. To investigate the role of pp42 MAP kinase in mediating c-Jun phosphorylation in vivo, we have treated U937 monocytic leukemia cells with a variety of pharmacological agents, including PMA, cycloheximide, AIF4, and okadaic acid. Although all of these agents stimulated c-Jun phosphorylation, cycloheximide and okadaic acid had no effect on pp42 MAP kinase phosphorylation, suggesting that MAP kinase activation was not necessary for c-Jun phosphorylation in vivo. Because dominant-negative RasAsn17 has been shown to block the effects of PMA on pp42 MAP kinase phosphorylation, we assessed its effect on c-Jun phosphorylation by cotransfection with a truncated c-Jun construct (c-Jun234). We found that c-Jun234 was expressed only in the cytosol and was inducibly phosphorylated with kinetics similar to those of endogenous nuclear c-Jun. Furthermore, we found that RasAsn17 had no effect on PMA-induced phosphorylation of c-Jun234. Because Ha-Ras requires isoprenylation for membrane binding, we examined the effect of the isoprenylation inhibitors lovastatin and perillic acid on PMA-induced c-Jun phosphorylation. Pretreatment of U937 cells with these agents had no effect on PMA-induced c-Jun or pp42 MAP kinase phosphorylation.(ABSTRACT TRUNCATED AT 250 WORDS)
Division of Hematology/Oncology, University of Alabama, Birmingham 35294.
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Franklin
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D002460:Cell Line; D003513:Cycloheximide / Q000494:pharmacology*; D004988:Ethers, Cyclic / Q000494:pharmacology*; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000187:drug effects / Q000502:physiology*; D011505:Protein-Tyrosine Kinases / Q000187:drug effects / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D011994:Recombinant Proteins / Q000187:drug effects; D015398:Signal Transduction / Q000502:physiology*
D004988:Ethers, Cyclic; D011518:Proto-Oncogene Proteins; D016755:Proto-Oncogene Proteins c-jun; D011994:Recombinant Proteins; D019319:Okadaic Acid; D003513:Cycloheximide; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D010749:Phosphoprotein Phosphatases