Extracellular signal-regulated kinases phosphorylate 5-lipoxygenase and stimulate 5-lipoxygenase product formation in leukocytes.
5-lipoxygenase (5-LO) is the key enzyme in the biosynthesis of proinflammatory leukotrienes. Here, we demonstrate that extracellular signal-regulated kinases (ERKs) can phosphorylate 5-LO in vitro. Efficient phosphorylation required the presence of unsaturated fatty acids and was abolished when Ser-663 was mutated to alanine. In intact HeLa cells stimulated with arachidonic acid (AA), impaired 5-LO product formation was evident in cells expressing the S663A-5-LO mutant compared with cells expressing wild-type 5-LO. For Mono Mac 6 cells, priming with phorbol myristate acetate (PMA) before stimulation with ionophore was required for ERK1/2 activation and efficient 5-LO phosphorylation, in parallel with substantial AA release and 5-LO product formation. Inhibition of PKC by GF109203x or MEK1/2 by U0126 (or PD98059) abolished the 5-LO up-regulation effects of PMA. In contrast, these inhibitors failed to suppress 5-LO product formation induced by stimuli such as AA plus ionophore, which apparently do not involve the ERK1/2 pathway. Based on inhibitor studies, ERKs are also involved in AA-stimulated 5-LO product formation in PMNL, whereas a role for ERKs is not apparent in 5-LO activation induced by ionophore or cell stress. Finally, the data suggest that ERKs and p38 MAPK-regulated MAPKAPKs can act in conjunction to stimulate 5-LO by phosphorylation.
Institute of Pharmaceutical Chemistry, University of Frankfurt, D-60439 Frankfurt, Germany. o.werz@pharmchem.uni-frankfurt.de
Oliver
O
Werz
Eva
E
Bürkert
Lutz
L
Fischer
Dagmar
D
Szellas
David
D
Dishart
Bengt
B
Samuelsson
Olof
O
Rådmark
Dieter
D
Steinhilber
D001094:Arachidonate 5-Lipoxygenase / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D015289:Leukotrienes / Q000096:biosynthesis*; D019950:Mitogen-Activated Protein Kinase 1 / Q000502:physiology; D020928:Mitogen-Activated Protein Kinases / Q000502:physiology*; D008954:Models, Biological; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D000099190:MAP-Kinase-Activated Kinase 2
D001094:Arachidonate 5-Lipoxygenase; D005231:Fatty Acids, Unsaturated; D047908:Intracellular Signaling Peptides and Proteins; D015289:Leukotrienes; D019950:Mitogen-Activated Protein Kinase 1; D020928:Mitogen-Activated Protein Kinases; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D000099190:MAP-Kinase-Activated Kinase 2