Identification of human prostaglandin E synthase: A microsomal, glutathione-dependent, inducible enzyme, constituting a potential novel drug target

Abstract

Human prostaglandin (PG) E synthase (EC 5.3.99.3) is a member of a recently recognized protein superfamily consisting of membrane associated proteins involved in eicosanoid and glutathione metabolism (the MAPEG family). Previous designations of the protein are PIG12 and MGST1-L1. PGE synthase was expressed in Escherichia coli, and both cytosolic and membrane fractions were prepared. Western blot analysis specifically detected a 15-to 16-kDa protein in the membrane fraction. Both fractions were incubated with prostaglandin H 2 in the presence or absence of reduced glutathione. The membrane but not the cytosolic fraction was found to possess high glutathione-dependent PGE synthase activity (0.25 mol͞ min͞mg). The human tissue distribution was analyzed by Northern blot analysis. High expression of PGE synthase mRNA was detected in A549 and HeLa cancer cell lines. Intermediate level of expression was demonstrated in placenta, prostate, testis, mammary gland, and bladder whereas low mRNA expression was observed in several other tissues. A549 cells have been used as a model system to study cyclooxygenase-2 induction by IL-1␤. If A549 cells were grown in the presence of IL-1␤, a significant induction of the PGE synthase was observed by Western blot analysis. Also, Western blot analysis specifically detected a 16-kDa protein in sheep seminal vesicles. In summary, we have identified a human membrane bound PGE synthase. The enzyme activity is glutathione-dependent, and the protein expression is induced by the proinflammatory cytokine IL-1␤. PGE synthase is a potential novel target for drug development.Prostaglandin endoperoxide H 2 (PGH 2 ) is formed from arachidonic acid by the action of cyclooxygenases (cox) -1 or -2. cox-1 is constitutively expressed in many cells and tissues such as platelets, endothelium, stomach, and kidney whereas the cox-2 protein can be induced by proinflammatory cytokines like IL-1␤ at sites of inflammation (for recent reviews on cox see refs. 1-3). Downstream of the cyclooxygenases, the product PGH 2 can be further metabolized into the various physiologically important eicosanoids: e.g., PGF 2␣ , PGE 2 , PGD 2 , PGI 2 (prostacyclin), and thromboxane A 2 (4).The mechanism for the biosynthesis of PGE 1 and PGF 1␣ (formed by using dihomo-␥-linolenic acid instead of arachidonic acid) (5) by sheep vesicular glands was postulated to proceed via a cyclic endoperoxide (6) later designated PGH 2 (7-9). In short, the reactions catalyzed by cyclooxygenase start by stereospecific abstraction of a hydrogen atom from carbon 13 of arachidonic acid. The resulting carbon radical reacts with molecular oxygen followed by the formation of the 9,11-endoperoxide and the bond between C-8 and C-12. Thereafter, a second molecule of oxygen is incorporated at C-15 followed by reduction to a hydroperoxy group, and PGG 2 is formed. This hydroperoxy group can subsequently be reduced by the peroxidase activity of the cyclooxygenase (in the presence of a reducing agent: e.g., glutathione) thus forming PGH 2 . The enzy...

Bradykinin potentiates cytokine‐induced prostaglandin biosynthesis in osteoblasts by enhanced expression of cyclooxygenase 2, resulting in increased RANKL expression

“…Moreover, the molecular mechanisms by which kinin receptors interact with IL-1 receptors, thus causing potentiation of PG formation and bone resorption, are not known. The potential mechanisms may include effects on cyclooxygenase 2 (COX-2) (PGH synthase 2), by which the conversion of arachidonic acid to the endoperoxide PGG 2 is catalyzed and followed by a reduction of PGG 2 to PGH 2 , and/or effects on the recently discovered PGE synthase (PGES), in which PGH 2 is converted to PGE 2 (20,21). Several forms of PGES seem to exist, including microsomal inducible PGES-1 (mPGES-1) as well as mPGES-2 and cytosolic PGES (cPGES) (22,23).…”

Expression of microsomal prostaglandin E synthase 1 in rheumatoid arthritis synovium

“…An inducible PGES has been identified as a 17-kd, glutathione-dependent integral membrane protein and is referred to as microsomal PGES-1 (mPGES-1) (3,4). Microsomal PGES-1 is induced by proinflammatory agents and is down-regulated by glucocorticoids in various cells (3,5,6).…”

“…Microsomal PGES-1 is induced by proinflammatory agents and is down-regulated by glucocorticoids in various cells (3,5,6). In contrast to the inducible mPGES-1, another mPGES (mPGES-2) is constitutively expressed in many cells without any further induction during inflammation (7).…”

La mPGES-1 : elle nous rend malades !

“…Dans le cas de la PGE2, ce sont les prostaglandine E synthases (PGES) qui permettent l'isomérisation de la PGH2 en PGE2. Alors que l'activité PGES est décrite depuis les années 1970, l'identification des enzymes est relativement récente [6]. Actuellement les gènes de trois isoformes des PGES ont été clonés : ceux de la PGES cytosolique (cPGES) et des PGES microsomales de type 1 et 2 (mPGES-1 et -2).…”

“…En particulier, la protéine d'échafaudage CARMA1 (caspase recruitment domain-containing membrane-associated guanylate kinase protein-1), et l'hétérodimère BCL10/ MALT1 (mucosa-associated lymphoid tissue) (complexe CBM) émergent comme des acteurs essentiels de la machinerie NF-κB ( Figure 1A) [5][6][7]. La protéine kinase C (PKC) θ ou β, dans les cellules T et B, respectivement, recrute et phosphoryle CARMA1 dans les microdomaines des radeaux lipidiques.…”