Abstract
5-Lipoxygenase activating protein (FLAP) functions as a facilitator of 5-lipoxygenase (5-LOX) activity. However, on the basis of the induction of apoptosis by the FLAP inhibitor MK886 in cells lacking 5-LOX, it is possible that this fatty acid-binding protein has other activities. This study was designed to examine potential roles of FLAP in apoptosis and cell proliferation. Overexpression of FLAP protein (2.2-fold) was achieved by stable transfection of IL-3-dependent murine prolymphoid progenitor cells (FL5.12) with a construct expressing the cDNA under a CMV promoter. The overexpressed protein was localized to nuclear membranes as with endogenous FLAP. The initial growth rate of FLAP-transfected cells was greater than that of control cells. After 48 h, when cell density had increased, the growth rate of FLAP-transfected cells declined substantially and there and there was a decrease in viability relative to control transfected cells. The FLAP-transfected cells were also more susceptible to withdrawal of IL-3 than were control cells. There was, however, no difference between FLAP and control cells in their susceptibility to MK886, NDGA, or etoposide during the log growth phase. Overexpression of FLAP did not alter Bcl-xL protein expression, but did decrease Bax protein and somewhat increased COX-1 and COX-2 mRNA levels. The failure of increased FLAP to alter susceptibility to MK886 provides further support to the concept that this agent induces apoptosis by mechanisms unrelated to FLAP. The data also suggest that FLAP can affect cell proliferation.
Similar content being viewed by others
References
Anderson KM, Levin J, Jajeh A, Seed T, Harris JE. Induction of apoptosis in blood cells from a patient with acute myelogenous leukemia by SC41661A, a selective inhibitor of 5-lipoxygenase. Prostaglandins Leukot Essen Fatty Acids 1993;48:323-6.
Anderson KM, Seed T, Plate JMD, Jajeh A, Meng J, Harris JE. Selective inhibitors of 5-lipoxygenase reduce CML blast cell proliferation and induce limited differentiation and apoptosis. Leuk Res. 1995;19:789-801.
Anderson KM, Seed T, Jajeh A, et al. An in vivo inhibitor of 5-lipoxygenase, MK886, at micromolar concentration induces apoptosis in U937 and CML cells. Anticancer Res. 1996;16:2589-600.
Avis IM, Jett M, Boyle T, et al. Growth control of lung cancer by interruption of 5-lipoxygenase-mediated growth factor signaling. J Clin Invest. 1996;97:806-13.
Battu S, Beneytout JL, Pairet M, Rigaud M. Cyclooxygenase-2 up-regulation after FLAP transfection in human adenocarcinoma cell line HT29 cl.19A. FEBS Lett. 1998;437:49-55.
Biswal SS, Datta K, Shaw SD, Feng X, Robertson JD, Kehrer JP. Glutathione oxidation and mitochondrial depolarization as mechanisms of nordihydroguaiaretic acid-induced apoptosis in lipoxygenase-deficient FL5.12 cells. Toxicol Sci. 2000;53:77-83.
Biswal SS, Datta K, Kehrer JP. Association between bcl-xL and 5-lipoxygenase activating protein (FLAP) levels in IL-3-dependent FL5.12 cells. Toxicology. 2001;160:97-103.
Buyn T, Dudeja P, Harris JE, et al. A 5-lipoxygenase inhibitor at micromolar concentration raises intracellular calcium in U937 cells prior to their physiologic cell death. Prostaglandins Leukot Essen Fatty Acids. 1997;56:69-77.
Claesson H-K, Jakobsson P-J, Steinhilber D, Odlander B, Samuelsson B. Expression of 5-lipoxygenase and biosynthesis of leukotriene b4 in human monomorphonuclear leukocytes J Lipid Mediat. 1993;6:15-22.
Coe NB, Bernlohr DA. Physiological properties and functions of intracellular fatty acid-binding proteins. Biochim Biophys Acta 1998;1391:287-306.
Datta K, Biswal SS, Xu J, Towndrow KM, Feng X, Kehrer JP. A relationship between 5-lipoxygenase activating protein (FLAP) and bcl-xL expression in murine pro B-lymphocytic FL5.12 cells. J Biol Chem. 1998;273:28163-9.
Datta K, Biswal SS, Kehrer JP. The 5-lipoxygenase activating protein (FLAP) inhibitor, MK886, induces apoptosis independent of FLAP. Biochem J. 1999;340:371-5.
Datta K, Kern JC, Biswal SS, Kehrer JP. Proteolysis and the loss of bcl-xL in FL5.12 cells undergoing apoptosis induced by MK886. Toxicol Appl Pharmacol. 2001;174:273-81.
el Makhour-Hojeij Y, Baclet MC, Chable-Rabinovitch H, Beneytout JL, Cook J. Expression of 5-lipoxygenase in lymphoblastoid B and T cells. Prostaglandins. 1994;48:21-9.
Ghosh J, Myers CE. Inhibition of arachidonate 5-lipoxygenase triggers massive apoptosis in human prostate cancer cells. Proc Natl Acad Sci USA 1998;95:13182-7.
Glatz JFC, Vork MM, Cistola DP, van der Vusse GJ. Cytoplasmic fatty acid-binding protein: significance for intracellular transport of fatty acids and putative role in signal transduction pathways. Prostaglandins Leukot Essent Fatty Acids. 1993;48:33-41.
Glatz JFC, Borchers T, Spener F, Van der Vusse GJ. Fatty acids in cell signalling: modulation by lipid binding proteins. Prostaglandins Leukot Essent Fatty Acids. 1995;52:121-7.
Glatz JFC, van Nieuwenhoven FA, Luiken JJFP, Schaap FG, van der Vusse GJ. Role of membrane-associated and cytoplasmic fatty acid binding proteins in cellular fatty acid metabolism. Prostaglandins Leukot Essent Fatty Acids. 1997;57:373-8.
Gugliucci A, Ranzato L, Scorrano L, et al. Mitochondria are direct targets of the lipoxygenase inhibitor MK886. A strategy for cell killing by combined treatment with MK886 and cyclooxygenase inhibitors. J Biol Chem. 2002;277:31789-95.
Kargman S, Rousseau P, Reid GK, et al. Leukotriene synthesis in U937 cells expressing recombinant 5-lipoxygenase. J Lipid Mediat. 1993;7:31-45.
Koller M, Wachtler P, David A, Muhr G, Konig W. Arachidonic acid induces DNA-fragmentation in human polymorphonuclear neutrophil granulocytes. Inflammation. 1997;21:463-74.
La E, Kern JC, Atarod EB, Kehrer JP. Fatty acid release and oxidation in lipoxygenase inhibitor-induced apoptosis. Toxicol Lett. 2003;138:193-203.
Lee JC, Hapel AJ, Ihle JN. Constitutive production of a unique lymphokine (IL 3) by the WEHI-3 cell line. J Immunol. 1982;128:2393-8.
Penzo D, Tagliapietra C, Colonna R, Petronilli V, Bernardi P. Effects of fatty acids on mitochondria: implications for cell death. Biochim Biophys Acta. 2002;1555:160-5.
Potter H. Introduction of DNA into mammalian cells-transfection by electroportion. In: Current protocols in molecular biology. New York: John Wiley; 1996; section 9.3.
Ramakrishnan N, Kalinich JF, McClain DE. Ebselen inhibition of apoptosis by reduction of peroxides. Biochem Pharmacol. 1996;51:1443-51.
Sandstrom PA, Tebbey PW, Van Cleave S, Buttke TM. Lipid hydroperoxides induce apoptosis in T cells displaying a HIV-associated glutathione peroxidase deficiency. J Biol Chem. 1994;269:798-801.
Sandstrom PA, Pardi D, Tebbey PW, et al. Lipid hydroperoxide-induced apoptosis: lack of inhibition by Bcl-2 over-expression. FEBS Lett. 1995;365:66-70.
Scorrano L, Penzo D, Petronilli V, Pagano F, Bernardi P. Arachidonic acid causes cell death through the mitochondrial permeability transition. Implications for tumor necrosis factor-a apoptotic signaling. J Biol Chem. 2001;276:12035-40.
Tang DG, Chen YQ, Honn KV. Arachidonate lipoxygenases as essential regulators of cell survival and apoptosis. Proc Natl Acad Sci USA 1996;93:5241-6.
Tang DG, La E, Kern J, Kehrer JP. Fatty acid oxidation and signaling in apoptosis. Biol Chem. 2002;383:425-42.
Vickers PJ. 5-Lipoxygenase activating protein (FLAP). J Lipid Mediat Cell Signal. 1995;12:185-94.
Woods JW, Coffey MJ, Brock TG, Singer II, Peters-Golden M. 5-Lipoxygenase is located in the euchromatin of the nucleus in resting human alveolar macrophages and translocates to the nuclear envelope upon cell activation. J Clin Invest. 1995;95:2035-46.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Wu, X., Biswal, S. & Kehrer, J. Roles of 5-lipoxygenase activating protein in cell proliferation and apoptosis. Cell Biol Toxicol 19, 135–143 (2003). https://doi.org/10.1023/A:1024789810277
Issue Date:
DOI: https://doi.org/10.1023/A:1024789810277