Extracellular signal‐regulated kinases phosphorylate 5lipoxygenase and stimulate 5‐lipoxygenase product formation in leukocytes

Werz, Oliver; Bürkert, Eva; Fischer, Lutz; Szellas, Dagmar; Dishart, David; Samuelsson, Bengt; Rådmark, Olof; Steinhilber, Dieter

doi:10.1096/fj.01-0909fje

Cited by 132 publications

(136 citation statements)

References 54 publications

Supporting

Mentioning

124

Contrasting

Order By: Relevance

“…The mechanisms involved here are still not fully understood, but there is evidence for cells with nuclear 5-LO to produce more LTB 4 than the same cells with cytoplasmic 5-LO (26). Phosphorylation by protein kinase (mitogen-activated protein) A on Ser523 can decrease 5-LO activity, whereas phosphorylation on Ser271 by MAP kinase (mitogen-activated protein) activated protein 2 and on Ser663 by extracellular signalregulated kinase can increase it (27)(28)(29).…”

Section: Regulation Of Lt Synthesismentioning

confidence: 99%

Leukotriene pathway genetics and pharmacogenetics in allergy

Duroudier

Tulah

Sayers

2009

Allergy

102

View full text Add to dashboard Cite

Leukotrienes (LT) are a family of potent eicosanoid lipid mediators with central importance in disease processes such as inflammation and proliferation most notably observed in allergic conditions like asthma (1). There are two general classes of LT according to the presence of a cysteine residue in their amino acid chain: the cysteinyl leukotrienes (CysLTs) including LTC 4 , LTD 4 and LTE 4 and the dihydroxyleukotriene LTB 4 (2). The 5-lipoxygenase pathwayLeukotrienes are produced in a multi-step enzyme pathway called the 5-lipoxygenase (5-LO) pathway, which is active in leucocytes such as neutrophils, eosinophils, mast cells and monocytes (Fig. 1). Arachidonic acid is the precursor for LT synthesis and is hydrolysed from the plasma membrane by cytosolic phospholipase A 2 (and other isoforms) (3, 4) in a calcium-dependent process (5). Upon activation, the rate-limiting enzyme 5-LO oxygenates first free arachidonic acid into the unstable intermediate 5-hydroperoxyeicosatetraenoic acid (5-HPETE) which is then either hydrolysed to 5-hydroxyeicosatetraenoic acid (5-HETE) or transformed into the unstable epoxide leukotriene A 4 (LTA 4 ) by forming a conjugated triene system through dehydration (6). 5-LO translocates from either the nucleus (in macrophages) or cyotosol (in neutrophils) to the nuclear envelope in response to cell activation (7,8). This movement occurs as 5-LO is dependent on a 5-LO activating protein (FLAP) for its function. FLAP remains associated with the nuclear membrane (9) and acts as a Ôtransfer proteinÕ presenting arachidonic acid to 5-LO, a system which allows the favourable conversion of 5-HPETE to LTA 4 compared to 5-HETE (10). Both 5-LO and FLAP are expressed in cells of myeloid lineage (11) restricting the pathway to these cell types. LTA 4 can be further metabolised to the cysteinyl leukotrienes (CysLTs) or LTB 4 . The specific glutathione S-transferase leukotriene C 4 synthase (LTC 4 S) conjugates LTA 4 to form LTC 4 which can then be rapidly converted to LTD 4 by a gammaglutamyl transpeptidase and to LTE 4 by a dipeptidase once exported out the cell by membrane transport proteins such as multi-drug related protein 1 (MRP1) (12-15). A specific zinc metallohydrolase, LTA 4 hydrolase (LTA 4 H) is responsible for the conversion of LTA 4 to LTB 4 (16). CysLTs and LTB 4 act on different G-protein coupled receptors (GPCRs). Each bind to at least two different receptors: CysLTs to CYSLTR1 and CYSLTR2 and LTB 4 to LTB 4 R1 and LTB 4 R2 (or BLT1 and BLT2) (17).Leukotrienes (LT) are biologically active lipid mediators known to be involved in allergic inflammation. Leukotrienes have been shown to mediate diverse features of allergic conditions including inflammatory cell chemotaxis/activation and smooth muscle contraction. Cysteinyl leukotrienes (LTC 4 , LTD 4 and, LTE 4 ) and the dihydroxy leukotriene LTB 4 are generated by a series of enzymes/ proteins constituting the LT synthetic pathway or 5-lipoxygenase (5-LO) pathway. Their function is mediated by interacting with multiple receptors. Leukotr...

show abstract

Section: Regulation Of Lt Synthesismentioning

confidence: 99%

Leukotriene pathway genetics and pharmacogenetics in allergy

Duroudier

Tulah

Sayers

2009

Allergy

102

View full text Add to dashboard Cite

show abstract

“…Recently, 5-LO kinases, namely the p38 MAPK-regulated MAP-KAPK-2/3, the ERK1/2, CaMKII and PKA [54], [55], [56], [57] the respective upstream signaling pathways could block cellular 5-LO product formation by such routes as well. In fact, the flavonoid quercetin 11 that suppresses 5-LO product synthesis also blocks the ERK1/2 and p38 MAPK pathway [58], [59], and plantderived tyrosine kinase inhibitors such as genistein reduced 5-LO activation in HL60 cells [47].…”

Section: Activation Pathways Of 5-lo In the Cellmentioning

confidence: 99%

Inhibition of 5-Lipoxygenase Product Synthesis by Natural Compounds of Plant Origin

Werz¹

2007

Planta Med

Self Cite

155

135

View full text Add to dashboard Cite

The biosynthesis of leukotrienes (LTs) is initiated by the transformation of free arachidonic acid to LTA 4 by 5-lipoxygenase (5-LO). Subsequent enzymatic conversion of LTA 4 yields LTB 4 and the cysteinyl-LTs C 4 , D 4 and E 4 . LTs have prominent functions in pathophysiology and are connected to numerous disorders including bronchial asthma, allergic rhinitis, inflammatory bowel and skin diseases, rheumatoid arthritis, cancer, osteoporosis and cardiovascular diseases. Pharmacological and genetic interruption of the 5-LO pathway or blockade of LT receptors, serving as means for intervention with LTs, may be of therapeutic value for certain related disorders. Natural or plant-derived substances were among the first 5-LO inhibitors identified in the early 1980 s. To date, a huge number of diverse plant-derived compounds have been reported to interfere with 5-LO product synthesis. However, many investigations have addressed the efficacy of a given compound solely in cellular test systems and analysis of direct interference with 5-LO has been neglected. In the first part of this review, the biology and molecular pharmacology of the 5-LO pathway is summarized in order to understand its overall regulation and complexity as well as to comprehend the possible points of attack of compounds that eventually lead to inhibition of 5-LO product formation in intact cells. In the second part, natural compounds that interfere with 5-LO product formation are compiled and grouped into structural classes, and the underlying molecular mechanisms and structureactivity relationships are discussed.

show abstract

“…With respect to PTMs of 5-LO, three phosphorylation sites for 5-LO have been described: Ser271, identified as mitogen-activated protein kinase activated protein kinase 2 (MK2) target site [5,6]; Ser663, described as the target site for extracellular signal-regulated kinase 2 (Erk2) [7]; and Ser523, characterized as a protein kinase A (PKA) target structure [8]. All three phosphorylations have been correlated to the regulation of 5-LO activity.…”

mentioning

confidence: 99%

“…The exact localization of the phosphorylation sites has been deduced by mutational analysis and in vitro phosphorylation assays with radioisotopically labeled ATP. However, they have never been verified on a molecular level by MS [5][6][7][8], which most probably reflects the considerable difficulties regarding purification of the enzyme from biological samples in a quality and quantity suitable for MS analysis, as well as the challenges regarding adequate sequence coverage by MS.…”

mentioning

confidence: 99%

Analysis of 5‐lipoxygenase phosphorylation on molecular level by MALDI‐MS

et al. 2014

Self Cite

View full text Add to dashboard Cite

The enzyme 5-lipoxygenase (5-LO) catalyzes the first reactions in the biosynthesis of leukotrienes, powerful lipid mediators that are involved in several physiological and pathological processes. 5-LO activity is tightly regulated by several factors, including post translational modifications (PTMs). Phosphorylations of 5-LO by the kinases extracellular signal-regulated kinase 2 (Erk2), mitogen-activated protein kinase activated protein kinase 2 (MK2) and protein kinase A (PKA) have been described to regulate 5-LO activity. Furthermore, 5-LO phosphorylation is considered a determinant of drug candidate potency. However, no evidence on a molecular level, as can be provided by MS, has as yet been presented for these PTMs. Here, we employ a workflow including different proteolytic cleavages and phosphopeptide enrichment for detection of 5-LO phosphorylation by MALDI-MS. Proof for the known phosphorylation sites of MK2 (Ser271) and PKA (Ser523) was provided by MS after in vitro phosphorylation, but not for the postulated Erk2 site (Ser663). Detection limits have been determined for all three sites. Moreover, we identified novel tyrosine kinase target sites within 5-LO using in silico and in vitro methods. Tyr42, Tyr53 and either Tyr94 or Tyr445 were phosphorylated by the Src kinases Fgr, hematopoietic cell kinase (HCK) and Yes. To analyze the phosphorylation state in the cellular context, we created stably 5-LO-transduced Mono Mac 6 cells. Here, we only detected phospho-Ser271 by MS, whereas immunoblot analysis indicated tyrosine phosphorylation, phospho-Ser271 and phospho-Ser663. Unexpectedly, phospho-Ser271 occurred independent of cell stimulation. Taken together, we describe a method for the molecular analysis of 5-LO phosphorylation, provide insights regarding the occurrence of known phosphorylation sites partly in contrast to earlier studies and present first evidence on novel phosphosites.

show abstract

Extracellular signal‐regulated kinases phosphorylate 5lipoxygenase and stimulate 5‐lipoxygenase product formation in leukocytes

Cited by 132 publications

References 54 publications

Leukotriene pathway genetics and pharmacogenetics in allergy

Leukotriene pathway genetics and pharmacogenetics in allergy

Inhibition of 5-Lipoxygenase Product Synthesis by Natural Compounds of Plant Origin

Analysis of 5‐lipoxygenase phosphorylation on molecular level by MALDI‐MS

Contact Info

Product

Resources

About