Phosphorylation of serine 271 on 5-lipoxygenase and its role in nuclear export.
The enzyme 5-lipoxygenase (5-LO) initiates the biosynthesis of leukotrienes, inflammatory mediators involved in immune diseases and defense. The subcellular localization of 5-LO is regulated, with nuclear import commonly leading to increased leukotriene production. We report here that 5-LO is constitutively phosphorylated on Ser-271 in transfected NIH 3T3 cells. This residue is nested in a classical nuclear export sequence, and phosphorylated Ser-271 5-LO was exclusively found in the nucleus by immunofluorescence and by fractionation techniques. Mutation of Ser-271 to Ala allowed nuclear export of 5-LO that was blocked by the specific nuclear export inhibitor leptomycin b, suggesting that phosphorylation of Ser-271 serves to interfere with exportin-1-mediated nuclear export. Consistent with previous reports that purified 5-LO can be phosphorylated on Ser-271 in vitro by MAPK-activated protein kinase 2, the nuclear export of 5-LO was increased by either treatment with the p38 inhibitor SB 203,580 or co-expression of a kinase-deficient p38 MAPK. Nuclear export of 5-LO can also be induced by KN-93, an inhibitor of Ca2+/calmodulin-dependent kinase II, and the effects of SB 203,580 plus KN-93 are additive. Finally, HeLa cells, which lack nuclear 5-LO, also lack constitutive phosphorylation of Ser-271. Taken together, these results indicate that the phosphorylation of Ser-271 serves to inhibit the nuclear export of 5-LO. This action works in concert with nuclear import, which is regulated by phosphorylation on Ser-523, to determine the subcellular distribution of 5-LO, which in turn regulates leukotriene biosynthesis.
Flamand
Nicolas
N
Department of Internal Medicine, Division of Pulmonary and Critical Care Medicine, University of Michigan Health System, Ann Arbor, Michigan 48109 and Centre de recherche de l'Hôpital Laval, Universitá Laval, Quábec City, QC G1V 4G5, Canada; Department of Internal Medicine, Division of Pulmonary and Critical Care Medicine, University of Michigan Health System, Ann Arbor, Michigan 48109 and Centre de recherche de l'Hôpital Laval, Universitá Laval, Quábec City, QC G1V 4G5, Canada. Electronic address: Nicolas.Flamand@crhl.ulaval.ca.
Luo
Ming
M
Department of Internal Medicine, Division of Pulmonary and Critical Care Medicine, University of Michigan Health System, Ann Arbor, Michigan 48109 and Centre de recherche de l'Hôpital Laval, Universitá Laval, Quábec City, QC G1V 4G5, Canada.
Peters-Golden
Marc
M
Department of Internal Medicine, Division of Pulmonary and Critical Care Medicine, University of Michigan Health System, Ann Arbor, Michigan 48109 and Centre de recherche de l'Hôpital Laval, Universitá Laval, Quábec City, QC G1V 4G5, Canada.
Brock
Thomas G
TG
Department of Internal Medicine, Division of Pulmonary and Critical Care Medicine, University of Michigan Health System, Ann Arbor, Michigan 48109 and Centre de recherche de l'Hôpital Laval, Universitá Laval, Quábec City, QC G1V 4G5, Canada.
D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D019943:Amino Acid Substitution; D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D001596:Benzylamines / Q000494:pharmacology; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D002467:Cell Nucleus / Q000201:enzymology* / Q000235:genetics; D006224:Cricetinae; D004285:Dogs; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D015289:Leukotrienes / Q000096:biosynthesis / Q000235:genetics; D048370:MAP Kinase Kinase 2 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D051379:Mice; D020125:Mutation, Missense; D041681:NIH 3T3 Cells; D049790:Nuclear Export Signals / Q000502:physiology*; D009893:Opossums; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D011817:Rabbits; D051381:Rats; D012694:Serine / Q000235:genetics / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D000096926:Benzenesulfonamides
D001094:Arachidonate 5-Lipoxygenase; D001596:Benzylamines; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D007093:Imidazoles; D015289:Leukotrienes; D048370:MAP Kinase Kinase 2; D049790:Nuclear Export Signals; D047428:Protein Kinase Inhibitors; D011725:Pyridines; D012694:Serine; D013449:Sulfonamides; D048051:p38 Mitogen-Activated Protein Kinases; C072105:KN 93; D000096926:Benzenesulfonamides; C093642:SB 203580