Thematic Review Series: Proteomics. An integrated omics analysis of eicosanoid biology
2009; Elsevier BV; Volume: 50; Issue: 6 Linguagem: Inglês
10.1194/jlr.r900004-jlr200
ISSN1539-7262
AutoresMatthew W. Buczynski, Darren S. Dumlao, Edward A. Dennis,
Tópico(s)Peroxisome Proliferator-Activated Receptors
ResumoEicosanoids have been implicated in a vast number of devastating inflammatory conditions, including arthritis, atherosclerosis, pain, and cancer. Currently, over a hundred different eicosanoids have been identified, with many having potent bioactive signaling capacity. These lipid metabolites are synthesized de novo by at least 50 unique enzymes, many of which have been cloned and characterized. Due to the extensive characterization of eicosanoid biosynthetic pathways, this field provides a unique framework for integrating genomics, proteomics, and metabolomics toward the investigation of disease pathology. To facilitate a concerted systems biology approach, this review outlines the proteins implicated in eicosanoid biosynthesis and signaling in human, mouse, and rat. Applications of the extensive genomic and lipidomic research to date illustrate the questions in eicosanoid signaling that could be uniquely addressed by a thorough analysis of the entire eicosanoid proteome. Eicosanoids have been implicated in a vast number of devastating inflammatory conditions, including arthritis, atherosclerosis, pain, and cancer. Currently, over a hundred different eicosanoids have been identified, with many having potent bioactive signaling capacity. These lipid metabolites are synthesized de novo by at least 50 unique enzymes, many of which have been cloned and characterized. Due to the extensive characterization of eicosanoid biosynthetic pathways, this field provides a unique framework for integrating genomics, proteomics, and metabolomics toward the investigation of disease pathology. To facilitate a concerted systems biology approach, this review outlines the proteins implicated in eicosanoid biosynthesis and signaling in human, mouse, and rat. Applications of the extensive genomic and lipidomic research to date illustrate the questions in eicosanoid signaling that could be uniquely addressed by a thorough analysis of the entire eicosanoid proteome. Biological processes are comprised of numerous converging signals that concertedly create a coherent effect. Any individual signaling element may elicit a physiological response, yet its loss is not fatal to the organism. In fact, a wide range of genetic variation can be observed within individual members of a given species without leading to a dramatic loss of function. As these signals have redundant and emergent properties, it can be difficult to explain how a biological process works using a limited number of molecular indicators. For this reason, systems biology has emerged to address the question of how molecular biology works as an integrated process (1Aderem A. Systems biology: its practice and challenges.Cell. 2005; 121: 511-513Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar). Systems biology has advanced exponentially during the past two decades, with transcriptomics, proteomics, and metabolomics each playing an integral role. Each of these platforms brings its own unique advantages and limitations in facilitating the investigation of disease pathology. A transcriptomic approach can detect the upregulation and downregulation of important biosynthetic and signaling genes; however, gene changes often don't directly correlate with changes in protein levels (2de Godoy L.M. Olsen J.V. Cox J. Nielsen M.L. Hubner N.C. Frohlich F. Walther T.C. Mann M. Comprehensive mass-spectrometry-based proteome quantification of haploid versus diploid yeast.Nature. 2008; 455: 1251-1254Crossref PubMed Scopus (697) Google Scholar). Proteomic analyses can identify enzymes and posttranslational protein changes in a cell or tissue, but fall short of determining which particular enzymes actively produce metabolites under disease conditions; likewise, metabolic approaches successfully identify bioactive signaling molecules, but therapeutic intervention generally requires definitive protein targets. The eicosanoid class of signaling molecules highlights many of the issues facing comprehensive systems biology analyses. Eicosanoids comprise a class of bioactive lipid mediators derived from the metabolism of polyunsaturated fatty acids by cyclooxygenases (3Funk C.D. Prostaglandins and leukotrienes: advances in eicosanoid biology.Science. 2001; 294: 1871-1875Crossref PubMed Scopus (2762) Google Scholar, 4Simmons D.L. Botting R.M. Hla T. Cyclooxygenase isozymes: the biology of prostaglandin synthesis and inhibition.Pharmacol. Rev. 2004; 56: 387-437Crossref PubMed Scopus (1203) Google Scholar–5Smith W.L. DeWitt D.L. Garavito R.M. Cyclooxygenases: structural, cellular, and molecular biology.Annu. Rev. Biochem. 2000; 69: 145-182Crossref PubMed Scopus (2278) Google Scholar), lipoxygenases (3Funk C.D. Prostaglandins and leukotrienes: advances in eicosanoid biology.Science. 2001; 294: 1871-1875Crossref PubMed Scopus (2762) Google Scholar, 6Serhan C.N. Chiang N. Van Dyke T.E. Resolving inflammation: dual anti-inflammatory and pro-resolution lipid mediators.Nat. Rev. Immunol. 2008; 8: 349-361Crossref PubMed Scopus (1884) Google Scholar), cytochrome P450s, or nonenzymatic pathways (Fig. 1). Technically, “eicosanoids” refers to fatty acids containing 20 carbons (eicosa), but the field generally uses the term eicosanoids more broadly to also include similar metabolites of other polyunsaturated fatty acids. This lipid class has been intensely studied over the past 30 years because of its contribution to the inflammatory response in diseases such as arthritis and asthma, yet the majority of this work has focused on a select few genes, proteins, or metabolites within it. In many instances, the regulation of one arm has important regulatory implications for another arm of the eicosanoid biosynthetic pathway. Many of these enzymes produce multiple lipid products; likewise, many lipid products can be formed by a number of different enzymes acting in parallel or in concert (Fig. 2). For these reasons, a comprehensive systems biology approach would be invaluable for understanding and treating diseases implicating eicosanoid signaling.Fig. 2Major eicosanoid biosynthetic pathways. The metabolites of the major pathways are indicated in color: COX (purple), 5-LOX (orange), 15-LOX (green), 12-LOX (yellow), CYP epoxygenase (red), CYP ω-hydroxylase (cyan), and nonenzymatic oxidation (gray). The products of arachidonic acid metabolism are illustrated, but similar products can be formed from other fatty acids (e.g., linoleic acid, eicosapentenoic acid, and docosahexaenoic acid).View Large Image Figure ViewerDownload Hi-res image Download (PPT) Transcriptomic and lipidomic analyses of eicosanoid biosynthesis can be performed using existing methodologies. With the completion of the major mammalian genome projects, commercial gene array technology has rapidly progressed and allowed the generation of whole-genome data sets of mRNA changes to become a commonplace tool at many research institutions. This proliferation allowed academic research groups to focus on the more challenging task of interpreting this data and made transcriptomic analysis the gold standard in systems biology research. Due to its immense scope, metabolomics has not yet achieved this level of standardization in scientific practice. However, as a part of their goal to create the infrastructure to identify and quantify all lipid molecular species, the Lipid Metabolites and Pathways Strategies (LIPID MAPS) consortium has developed liquid chromatography tandem mass spectrometry methodology that comprehensively covers nearly all the known metabolites of this class (7Feng L. Prestwich G.D. Functional Lipidomics. Taylor & Francis, Boca Raton, FL2006Google Scholar, 8Schmelzer K. Fahy E. Subramaniam S. Dennis E.A. The lipid maps initiative in lipidomics.Methods Enzymol. 2007; 432: 171-183Crossref PubMed Scopus (99) Google Scholar). Specifically, a complete lipidomic analysis of eicosanoids is now available (9Buczynski M.W. Stephens D.L. Bowers-Gentry R.C. Grkovich A. Deems R.A. Dennis E.A. TLR-4 and sustained calcium agonists synergistically produce eicosanoids independent of protein synthesis in RAW264.7 cells.J. Biol. Chem. 2007; 282: 22834-22847Abstract Full Text Full Text PDF PubMed Scopus (76) Google Scholar), along with corresponding gene microarray data (www.lipidmaps.org). Thus, is it timely to consider a proteomic analysis that could complete an integrated picture of the entire eicosanoid signaling network. A quantitative proteomic analysis of the eicosanoid pathway has thus far lagged behind concomitant transcriptomic and lipidomic studies. These tandem mass spectrometry techniques can be divided into two philosophical approaches (10Domon B. Aebersold R. Mass spectrometry and protein analysis.Science. 2006; 312: 212-217Crossref PubMed Scopus (1460) Google Scholar). The classically practiced MS1 and MS2 proteomic scanning techniques have demonstrated the potential to rapidly identify thousands of proteins from a single sample, yet these often represent only a fraction of the total protein content of a cell or tissue, and the investigator has little control over which proteins make the cut (11Bantscheff M. Schirle M. Sweetman G. Rick J. Kuster B. Quantitative mass spectrometry in proteomics: a critical review.Anal. Bioanal. Chem. 2007; 389: 1017-1031Crossref PubMed Scopus (1144) Google Scholar). On the other hand, by measuring the levels of a selected population of proteins using multiple reaction monitoring, significantly lower limits of detection for can be achieved (12Keshishian H. Addona T. Burgess M. Kuhn E. Carr S.A. Quantitative, multiplexed assays for low abundance proteins in plasma by targeted mass spectrometry and stable isotope dilution.Mol. Cell. Proteomics. 2007; 6: 2212-2229Abstract Full Text Full Text PDF PubMed Scopus (552) Google Scholar). However, to create a method to investigate a selected proteome, one must first have a fundamental understanding of what proteins could be involved and how these proteins fit into the larger scheme. In the book Functional Lipidomics, Bowers-Gentry et al. (7Feng L. Prestwich G.D. Functional Lipidomics. Taylor & Francis, Boca Raton, FL2006Google Scholar) give a broad outline for using a lipidomic approach to classify and measure eicosanoids. To facilitate a concerted systems biology approach, this review provides a systematic overview of the proteins involved in eicosanoid biosynthesis and signaling. The genes and proteins responsible for these activities are listed in supplementary Tables I–VII online, provided one of the following pieces of evidence: 1) the gene has been expressed and characterized in vitro; 2) the purified protein has been sequenced and characterized in vitro; 3) the gene has been overexpressed (or for transcription factors, expressed with a reporter gene) and characterized in a cellular system; 4) loss of activity has been demonstrated in cells or tissue from gene knockout animals. Furthermore, as studies of human biology and disease commonly employ mouse or rat models as a surrogate, important differences between these species are highlighted. Arachidonic acid and other polyunsaturated fatty acids serve as the metabolic precursors for eicosanoid synthesis (13Schaloske R.H. Dennis E.A. The phospholipase A2 superfamily and its group numbering system.Biochim. Biophys. Acta. 2006; 1761: 1246-1259Crossref PubMed Scopus (670) Google Scholar, 14Six D.A. Dennis E.A. The expanding superfamily of phospholipase A(2) enzymes: classification and characterization.Biochim. Biophys. Acta. 2000; 1488: 1-19Crossref PubMed Scopus (1158) Google Scholar). Biologically, these molecules are generally not available in large quantities in the free acid form, but are stored at the sn-2 position on the glycerol backbone of membrane phospholipids. To be used for biosynthesis, phospholipase A2 (PLA2) liberates sn-2 fatty acids from phospholipids at the membrane interface. Phospholipase A2 represents a superfamily of at least 15 groups that have wide-ranging roles in biological processes. These enzymes can be considered as five types: cytosolic PLA2 (cPLA2), secreted PLA2 (sPLA2), calcium-independent PLA2, platelet-activating factor acetylhydrolase, and lysosomal PLA2; their classification and biological functions have been extensively reviewed (13Schaloske R.H. Dennis E.A. The phospholipase A2 superfamily and its group numbering system.Biochim. Biophys. Acta. 2006; 1761: 1246-1259Crossref PubMed Scopus (670) Google Scholar, 14Six D.A. Dennis E.A. The expanding superfamily of phospholipase A(2) enzymes: classification and characterization.Biochim. Biophys. Acta. 2000; 1488: 1-19Crossref PubMed Scopus (1158) Google Scholar–15Burke J.E. Dennis E.A. Phospholipase A2 structure/function, mechanism and signaling.J. Lipid Res. November, 14, 2008; (Epub ahead of print)doi: 10.1194/jlr.R800033-JLR200PubMed Google Scholar). Current literature strongly implicates the Group IVA cPLA2 as the chief enzyme involved in polyunsaturated fatty acid release for eicosanoid biosynthesis (16Uozumi N. Shimizu T. Roles for cytosolic phospholipase A2alpha as revealed by gene-targeted mice.Prostaglandins Other Lipid Mediat. 2002; 68–69: 59-69Crossref PubMed Scopus (72) Google Scholar, 17Ghosh M. Tucker D.E. Burchett S.A. Leslie C.C. Properties of the Group IV phospholipase A2 family.Prog. Lipid Res. 2006; 45: 487-510Crossref PubMed Scopus (288) Google Scholar), with sPLA2s potentially playing a supplementary role (see supplementary Table I). Bonventre et al. (18Bonventre J.V. Huang Z. Taheri M.R. O'Leary E. Li E. Moskowitz M.A. Sapirstein A. Reduced fertility and postischaemic brain injury in mice deficient in cytosolic phospholipase A2.Nature. 1997; 390: 622-625Crossref PubMed Scopus (730) Google Scholar) and Uozumi et al. (19Uozumi N. Kume K. Nagase T. Nakatani N. Ishii S. Tashiro F. Komagata Y. Maki K. Ikuta K. Ouchi Y. et al.Role of cytosolic phospholipase A2 in allergic response and parturition.Nature. 1997; 390: 618-622Crossref PubMed Scopus (614) Google Scholar) independently demonstrated that peritoneal macrophages from Group IVA PLA2 knockout mice were incapable of producing eicosanoids. More recently, Adler et al. (20Adler D.H. Cogan J.D. Phillips J.A. Schnetz-Boutaud N. Milne G.L. Iverson T. Stein J.A. Brenner D.A. Morrow J.D. Boutaud O. et al.Inherited human cPLA(2alpha) deficiency is associated with impaired eicosanoid biosynthesis, small intestinal ulceration, and platelet dysfunction.J. Clin. Invest. 2008; 118: 2121-2131PubMed Google Scholar) identified a patient with inherited Group IVA deficiency and determined that nearly all arachidonic acid used for eicosanoid production by platelets and circulating leukocytes was attributable to this enzyme. The Group IVA cPLA2 uses a catalytic Ser/Asp dyad to hydrolyze fatty acids and contains a C2 domain that facilitates calcium-dependent translocation from the cytosol to the membrane surface where it encounters phospholipid substrate. Overall, while cPLA2s have a broad range of homology between human, mouse, and rat, the Group IVA cPLA2 has 95% identity between these species. Other cPLA2 show between 54 and 82% identity in these species and have been less well-characterized to date (13Schaloske R.H. Dennis E.A. The phospholipase A2 superfamily and its group numbering system.Biochim. Biophys. Acta. 2006; 1761: 1246-1259Crossref PubMed Scopus (670) Google Scholar), yet may still play a supplementary role in eicosanoid biosynthesis. While Group IVA cytosolic PLA2 is the primary enzyme involved in eicosanoid biosynthesis, sPLA2s have been demonstrated to play a supplementary role, reviewed extensively by Lambeau and Gelb (21Lambeau G. Gelb M.H. Biochemistry and physiology of mammalian secreted phospholipases A2.Annu. Rev. Biochem. 2008; 77: 495-520Crossref PubMed Scopus (373) Google Scholar). In particular, the Group IIA, Group V, and Group X PLA2s have all been shown to modulate eicosanoid levels in mammalian cellular systems. However, the majority of these studies have been performed by either overexpression or exogenous supplementation of sPLA2, which complicates the investigation of its role in pathology. Future work using sPLA2 knockout mice may help further elucidate the role these enzymes play in eicosanoid biosynthesis during an inflammatory response. Prostaglandins (PGs) are bioactive signaling molecules derived from cyclooxygenase (COX) and subsequent PG synthase activity on arachidonic acid (4Simmons D.L. Botting R.M. Hla T. Cyclooxygenase isozymes: the biology of prostaglandin synthesis and inhibition.Pharmacol. Rev. 2004; 56: 387-437Crossref PubMed Scopus (1203) Google Scholar, 5Smith W.L. DeWitt D.L. Garavito R.M. Cyclooxygenases: structural, cellular, and molecular biology.Annu. Rev. Biochem. 2000; 69: 145-182Crossref PubMed Scopus (2278) Google Scholar, 22Rouzer C.A. Marnett L.J. Cyclooxygenases: structural and functional insights.J. Lipid Res. October 23, 2008; (Epub ahead of print.)doi 10.1194/jlr.R800042-JLR200PubMed Google Scholar) (see supplementary Table II). COXs contain two distinct active sites, a COX and peroxidase site, both of which use the same tyrosyl radical and heme-iron for catalysis. The COX site incorporates molecular O2 at the 11- and 15-carbon on arachidonic acid to form PGG2, which contains the following moieties: a five-member ring linked at C-8 and C-12, an endoperoxide bridge across C-9 and C-11, and a peroxide at C-15. The peroxidase site reduces the peroxide to a hydroxyl to form PGH2, the substrate for the various PG synthases. In addition to forming PGH2, arachidonic acid can situate in the active site pocket in a limited number of suboptimal conformations that incorporate a single molecular O2, forming trace amounts of either 11(R)-hydroxyeicosatetraenoic acid (HETE) or 15(S)-HETE. COX activity does not exhibit long-term stability, as the catalytic tyrosyl radical can be transferred to a nearby tyrosyl residue and cause “suicide inactivation” after ∼300 turnovers. Because the enzyme can perform only a limited number of reactions, it must be constantly reexpressed to generate metabolites. The details of these mechanisms have been reviewed extensively (5Smith W.L. DeWitt D.L. Garavito R.M. Cyclooxygenases: structural, cellular, and molecular biology.Annu. Rev. Biochem. 2000; 69: 145-182Crossref PubMed Scopus (2278) Google Scholar). COXs constitute two distinct genes, COX-1 and COX-2. These two isoforms exhibit ∼60% identity and have nearly identical active site residues (4Simmons D.L. Botting R.M. Hla T. Cyclooxygenase isozymes: the biology of prostaglandin synthesis and inhibition.Pharmacol. Rev. 2004; 56: 387-437Crossref PubMed Scopus (1203) Google Scholar). The most significant structural difference between these enzymes is an isoleucine to valine substitution, which results in a larger COX-2 active site pocket. This allows COX-2 to be more permissive in selecting substrates, and unlike COX-1, it can metabolize dihomo-γ-linolenic and eicosapentaenoic acid in addition to arachidonic acid. Pharmaceutical companies have also taken advantage of the larger COX-2 active site, creating isoform selective inhibitors such as Celecoxib (Celebrex™). While COX-1 and COX-2 have similar structural and catalytic features, these isozymes exhibit different expression patterns. COX-1 is constitutively expressed by most cell types and has been implicated in a number of homeostatic processes, including stomach acidity control, endometrial cycling, and renal function. In contrast, COX-2 expression is controlled by the pro-inflammatory transcription factor NF-κB and highly upregulated in response to infection, atherosclerosis, and a number of cancers. PGH2 can form a number of different bioactive products through the action of PG synthases. This includes a number of important signaling molecules, including PGI2 (also known as prostacyclin), thromboxane A2, PGE2 (also known as dinoprostone), PGD2, and PGF2α. Following the PG abbreviation (Fig. 3), the nomenclature highlights the two important structural features: the components of the five-member prostane ring, as denoted by a letter A–K, and the number of double bonds, denoted by a subscript number (23Nelson N.A. Prostaglandin nomenclature.J. Med. Chem. 1974; 17: 911-918Crossref PubMed Google Scholar). The lone exception is thromboxane, which has a six-member oxane ring, and is abbreviated TX. PGI2 was first identified in 1976 (24Whittaker N. Bunting S. Salmon J. Moncada S. Vane J.R. Johnson R.A. Morton D.R. Kinner J.H. Gorman R.R. McGuire J.C. et al.The chemical structure of prostaglandin X (prostacyclin).Prostaglandins. 1976; 12: 915-928Crossref PubMed Scopus (584) Google Scholar) and is formed by the prostacyclin synthase, a member of the cytochrome P450 monooxygenase superfamily (25Wu K.K. Liou J.Y. Cellular and molecular biology of prostacyclin synthase.Biochem. Biophys. Res. Commun. 2005; 338: 45-52Crossref PubMed Scopus (72) Google Scholar). Structural elucidation of the human prostacyclin synthase was not obtained until 2006 (26Chiang C.W. Yeh H.C. Wang L.H. Chan N.L. Crystal structure of the human prostacyclin synthase.J. Mol. Biol. 2006; 364: 266-274Crossref PubMed Scopus (54) Google Scholar), confirming the mechanism of PGH2 endoperoxide bridge rearrangement proposed by Hecker and Ullrich in 1989 (27Hecker M. Ullrich V. On the mechanism of prostacyclin and thromboxane A2 biosynthesis.J. Biol. Chem. 1989; 264: 141-150Abstract Full Text PDF PubMed Google Scholar). The active site heme-iron interacts with the C-11 oxygen, promoting the hemolytic cleavage of the endoperoxide bond and the formation of an ether linkage between C-9 and C-5. The PGI2 ring is highly labile and rapidly hydrolyzed to form the stable but biologically inactive 6-keto PGF1α, and because of this, PGI2 and 6-keto PGF1α are often used interchangeably in the literature. PGI2 binds the G-coupled protein receptor IP (28Katsuyama M. Sugimoto Y. Namba T. Irie A. Negishi M. Narumiya S. Ichikawa A. Cloning and expression of a cDNA for the human prostacyclin receptor.FEBS Lett. 1994; 344: 74-78Crossref PubMed Scopus (0) Google Scholar, 29Boie Y. Rushmore T.H. Darmon-Goodwin A. Grygorczyk R. Slipetz D.M. Metters K.M. Abramovitz M. Cloning and expression of a cDNA for the human prostanoid IP receptor.J. Biol. Chem. 1994; 269: 12173-12178Abstract Full Text PDF PubMed Google Scholar) as well as the transcription factors peroxisome proliferator-activated receptor (PPAR) α, PPARδ, and PPARγ (30Aubert J. Ailhaud G. Negrel R. Evidence for a novel regulatory pathway activated by (carba)prostacyclin in preadipose and adipose cells.FEBS Lett. 1996; 397: 117-121Crossref PubMed Scopus (40) Google Scholar, 31Kojo H. Fukagawa M. Tajima K. Suzuki A. Fujimura T. Aramori I. Hayashi K. Nishimura S. Evaluation of human peroxisome proliferator-activated receptor (PPAR) subtype selectivity of a variety of anti-inflammatory drugs based on a novel assay for PPAR delta(beta).J. Pharmacol. Sci. 2003; 93: 347-355Crossref PubMed Scopus (0) Google Scholar). Identified in 1975 (32Hamberg M. Svensson J. Samuelsson B. Thromboxanes: a new group of biologically active compounds derived from prostaglandin endoperoxides.Proc. Natl. Acad. Sci. USA. 1975; 72: 2994-2998Crossref PubMed Google Scholar), thromboxane A2 (TXA2) is the physiological counterbalancing signal to PGI2. Similar to PGI2, its biosynthesis is catalyzed by a member of the cytochrome P450 superfamily member, thromboxane A synthase (27Hecker M. Ullrich V. On the mechanism of prostacyclin and thromboxane A2 biosynthesis.J. Biol. Chem. 1989; 264: 141-150Abstract Full Text PDF PubMed Google Scholar, 33Kinsella B.T. O'Mahony D.J. Fitzgerald G.A. The human thromboxane A2 receptor alpha isoform (TP alpha) functionally couples to the G proteins Gq and G11 in vivo and is activated by the isoprostane 8-epi prostaglandin F2 alpha.J. Pharmacol. Exp. Ther. 1997; 281: 957-964PubMed Google Scholar). This enzyme facilitates rearrangement of the PGH2 endoperoxide bridge by a complimentary mechanism to prostacyclin synthase, interacting with the C-9 oxygen to promote endoperoxide bond cleavage. The C-11 oxygen radical initiates intramolecular rearrangement, resulting in either the formation of TXA2 or 12-hydroxyheptadecatrienoic acid. TXA2 contains an unstable ether linkage that is rapidly hydrolyzed in vivo under aqueous conditions to form the biologically inert TXB2. It also binds a specific G protein-coupled receptor (GPCR), termed the TP receptor, which culminates in increasing intracellular calcium concentrations (33Kinsella B.T. O'Mahony D.J. Fitzgerald G.A. The human thromboxane A2 receptor alpha isoform (TP alpha) functionally couples to the G proteins Gq and G11 in vivo and is activated by the isoprostane 8-epi prostaglandin F2 alpha.J. Pharmacol. Exp. Ther. 1997; 281: 957-964PubMed Google Scholar). PGE2 may be the best characterized signaling molecule within the eicosanoid class. A PubMed search for “PGE2” reveals that, since the identification and isolation of PGE2 from human seminal plasma in 1963 (34Samuelsson B. Isolation and identification of prostaglandins from human seminal plasma. 18. Prostaglandins and related factors.J. Biol. Chem. 1963; 238: 3229-3234Abstract Full Text PDF PubMed Google Scholar), well over 27,000 articles have been published studying this molecule. The effects of PGE2 have been implicated in innumerable biological processes and disease pathologies, including parturition, bronchodilation, pain signaling, innate and adaptive immune responses, cancer, arthritis, and atherosclerosis (4Simmons D.L. Botting R.M. Hla T. Cyclooxygenase isozymes: the biology of prostaglandin synthesis and inhibition.Pharmacol. Rev. 2004; 56: 387-437Crossref PubMed Scopus (1203) Google Scholar, 35Park J.Y. Pillinger M.H. Abramson S.B. Prostaglandin E2 synthesis and secretion: the role of PGE2 synthases.Clin. Immunol. 2006; 119: 229-240Crossref PubMed Google Scholar). The wide spectrum of PGE2 signaling can be partially attributed to differences in expression and function of its four known receptors, EP1-4. The EP receptors are all GPCRs, and their most potent natural ligand is PGE2. Functionally, EP1 and EP3 mediate their responses through calcium signaling, whereas EP2 and EP4 do so by increasing cAMP levels. Likewise, PGE2 synthesis occurs through three unique enzymes, named the cytosolic PGE synthase (cPGES), microsomal PGE synthase-1 (mPGES-1) and mPGES-2 (35Park J.Y. Pillinger M.H. Abramson S.B. Prostaglandin E2 synthesis and secretion: the role of PGE2 synthases.Clin. Immunol. 2006; 119: 229-240Crossref PubMed Google Scholar). Cytosolic PGE2 synthase was identified by Tanioka et al. (36Tanioka T. Nakatani Y. Semmyo N. Murakami M. Kudo I. Molecular identification of cytosolic prostaglandin E2 synthase that is functionally coupled with cyclooxygenase-1 in immediate prostaglandin E2 biosynthesis.J. Biol. Chem. 2000; 275: 32775-32782Abstract Full Text Full Text PDF PubMed Scopus (602) Google Scholar) in 2000 from rat brain and determined it to be identical to human p23. Previously, this protein was identified as a chaperone protein that interacts with heat shock protein-90, which upregulates cPGES activity in vitro. The catalytic activity of cPGES requires glutathione and the conserved Tyr-9 residue for activity, similar to other cytosolic glutathione S-transferases, including hematopoietic PGD synthase (35Park J.Y. Pillinger M.H. Abramson S.B. Prostaglandin E2 synthesis and secretion: the role of PGE2 synthases.Clin. Immunol. 2006; 119: 229-240Crossref PubMed Google Scholar, 37Murakami M. Nakatani Y. Tanioka T. Kudo I. Prostaglandin E synthase.Prostaglandins Other Lipid Mediat. 2002; 68–69: 383-399Crossref PubMed Scopus (216) Google Scholar). In this case, the glutathione thiol is expected to interact with the C-9 oxygen of PGH2, facilitating cleavage of the endoperoxide bridge and the formation of PGE2. The cloning and characterization of human mPGES-1 was reported by Jakobsson et al. in 1999 (38Jakobsson P.J. Thoren S. Morgenstern R. Samuelsson B. Identification of human prostaglandin E synthase: a microsomal, glutathione-dependent, inducible enzyme, constituting a potential novel drug target.Proc. Natl. Acad. Sci. USA. 1999; 96: 7220-7225Crossref PubMed Scopus (853) Google Scholar). As a member of the membrane-associated proteins in eicosanoid and glutathione (MAPEG) superfamily of transmembrane proteins, this enzyme requires glutathione for activity. While mPGES-1 has not yet been crystallized, Huang et al. (39Huang X. Yan W. Gao D. Tong M. Tai H.H. Zhan C.G. Structural and functional characterization of human microsomal prostaglandin E synthase-1 by computational modeling and site-directed mutagenesis.Bioorg. Med. Chem. 2006; 14: 3553-3562Crossref PubMed Scopus (0) Google Scholar) examined it using site-directed mutagenesis and computational modeling onto the microsomal glutathione S-transferase (mGST) structure, its nearest subfamily member. This method identified Arg-110 and Tyr-114 interactions with the C terminus of PGH2 and determined that glutathione thiol was positioned to catalyze PGE2 biosynthesis. The mechanism of PGE2 formation likely proceeds by a similar mechanism to cPGES and h-PGDS; however, the structural element activating glutathione remains undetermined. mPGES-1 expression is coregulated with COX-2, and both proteins colocalize to the endoplasmic reticulum (40Samuelsson B. Morgenstern R. Jakobsson P.J. Membrane prostaglandin E synthase-1: a novel therapeutic target.Pharmacol. Rev. 2007; 59: 207-224Crossref PubMed Scopus (408) Google Scholar); however, COX-2 resides in the lumen of the endoplasmic reticulum, whereas the mPGES-1 active site appears to face the cytosol. A thorough examination of mPGES-1 latur
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