Molecular Identification of Cytosolic Prostaglandin E2 Synthase That Is Functionally Coupled with Cyclooxygenase-1 in Immediate Prostaglandin E2Biosynthesis

Abstract

Here we report the molecular identification of cytosolic glutathione (GSH)-dependent prostaglandin (PG) E 2 synthase (cPGES), a terminal enzyme of the cyclooxygenase (COX)-1-mediated PGE 2 biosynthetic pathway. GSHdependent PGES activity in the cytosol of rat brains, but not of other tissues, increased 3-fold after lipopolysaccharide (LPS) challenge. Peptide microsequencing of purified enzyme revealed that it was identical to p23, which is reportedly the weakly bound component of the steroid hormone receptor/hsp90 complex. Recombinant p23 expressed in Escherichia coli and 293 cells exhibited all the features of PGES activity detected in rat brain cytosol. A tyrosine residue near the N terminus (Tyr 9 ), which is known to be critical for the activity of cytosolic GSH S-transferases, was essential for PGES activity. The expression of cPGES/p23 was constitutive and was unaltered by proinflammatory stimuli in various cells and tissues, except that it was increased significantly in rat brain after LPS treatment. cPGES/p23 was functionally linked with COX-1 in marked preference to COX-2 to produce PGE 2 from exogenous and endogenous arachidonic acid, the latter being supplied by cytosolic phospholipase A 2 in the immediate response. Thus, functional coupling between COX-1 and cPGES/p23 may contribute to production of the PGE 2 that plays a role in maintenance of tissue homeostasis. Biosynthesis of prostaglandin (PG)1 E 2 , the most common prostanoid with potent bioactivities, is regulated by three sequential steps of the cyclooxygenase (COX) pathway. Phospholipase A 2 (PLA 2 ) initiates this pathway by releasing arachidonic acid (AA) from membrane glycerophospholipids. Of more than 10 members of the PLA 2 family characterized to date, cytosolic PLA 2 (cPLA 2 ) and several secretory PLA 2 s are involved in supplying AA to either of the two COX isozymes, COX-1 and COX-2, depending upon the phases of cell activation (1-3). The constitutive COX-1 is mainly utilized in immediate PGE 2 biosynthesis, which occurs within several minutes after stimulation with Ca 2ϩ mobilizers, whereas the inducible COX-2 mediates the delayed PGE 2 biosynthesis, which lasts for several hours following proinflammatory stimuli. Although COX-1 and COX-2 have been reported to exhibit subtle differences in AA requirements in that COX-2 is favored over COX-1 at low AA concentrations (3-5) and subcellular localizations (6), their functional segregation in the PGE 2 biosynthetic response cannot be fully explained only by these aspects.The activity of PGES, which catalyzes conversion of COXderived PGH 2 to PGE 2 , has been detected in both cytosolic and microsomal fractions of various cells, and in most, if not all, cases it requires glutathione (GSH) for optimal activity (7-9). Although several groups have attempted to purify this critical enzyme to near homogeneity for the last 20 years (7-9), such trials have been unsuccessful. The PGES enzyme purified from human brain cytosol was identified as a GSH S-transferase (GST), which converts PGH 2 to PGE ...

FcεRI, but Not FcγR, Signals Induce Prostaglandin D2 and E2 Production from Basophils

“…[35][36][37] Expression of mPGES1 is markedly induced by pro-inflammatory stimulation in various tissues. 35 PGES1 is involved in the COX-2-mediated PGE2-biosynthetic pathway 35 ; mPGES2 is abundant in brain, heart, skeletal muscle, kidney, and liver 37 ; cPGES is distributed ubiquitously in the cytosol of various cells 36 ; and COX-1 contributes to PGE2 generation by PGES2 and cPGES.…”

“…35 PGES1 is involved in the COX-2-mediated PGE2-biosynthetic pathway 35 ; mPGES2 is abundant in brain, heart, skeletal muscle, kidney, and liver 37 ; cPGES is distributed ubiquitously in the cytosol of various cells 36 ; and COX-1 contributes to PGE2 generation by PGES2 and cPGES. 36,39,40 We immunohistochemically confirmed the presence of mPGES1 in primary basophils, but we were unable to assess the protein expression of the other PGES enzymes, because antibodies against mPGES2 and cPGES were not available in our laboratory. Nevertheless, transcripts encoding mPGES1, mPGES2, and cPGES were all detected in BMBAs.…”

“…PGES is classified into three types, glutathione-dependent membrane-bound PGES (mPGES1), glutathione-independent membrane-bound PGES (mPGES2), and cytosolic PGES (cPGES). [35][36][37] Hpgds transcripts that encode H-PGDS were readily detected in both BMBAs and BMMCs by RT-PCR. In contrast, Ptgds transcripts that encode L-PGDS were only detected in BMMCs ( Figure 5, A and B).…”

Expression of microsomal prostaglandin E synthase 1 in rheumatoid arthritis synovium

“…In contrast to the inducible mPGES-1, another mPGES (mPGES-2) is constitutively expressed in many cells without any further induction during inflammation (7). In addition, a cytosolic protein possessing PGES activity (cPGES) has been characterized and found to be constitutively expressed in various cells (8). Cytosolic PGES is not generally induced by proinflammatory stimuli except in the brain, where a minor increase has been reported (8).…”

“…In addition, a cytosolic protein possessing PGES activity (cPGES) has been characterized and found to be constitutively expressed in various cells (8). Cytosolic PGES is not generally induced by proinflammatory stimuli except in the brain, where a minor increase has been reported (8). It has been demonstrated that mPGES-1 functionally uses PGH 2 generated by COX-2 (5,6), but some evidence also exists for a role of COX-1 (9).…”