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Molecular characterization of the 8‐iso‐PGF 2 α interaction with the thromboxane A 2 receptor and its signaling mechanisms in human platelets
Author(s) -
Khasawneh Fadi T,
Huang JinSheng,
Tiruppathi Chinnaswamy,
Le Breton Guy C
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a427
Subject(s) - thromboxane receptor , eicosanoid , chemistry , isoprostanes , arachidonic acid , platelet , receptor , isoprostane , thromboxane , prostaglandin , signal transduction , prostaglandin d2 , thromboxane a2 , rhoa , prostaglandin h2 , platelet activation , prostanoid , biochemistry , medicine , oxidative stress , enzyme , lipid peroxidation
Isoprostanes are nonenzymatic products of arachidonic acid that are thought to be involved in the pathogenesis of different diseases. In spite of this involvement, however, their signal transduction mechanisms are unclear. The present study examined whether isoprostanes interact with known eicosanoid receptors, e.g. thromboxane A 2 (TP), prostaglandin F 2 α (FP), PGI 2 (IP), PGD 2 (DP) or PGE 2 (EP). It was found, that 8‐iso‐PGF 2 α mobilized calcium and bound to cells expressing the wild‐type TP (K d =57 nM). We next defined the amino acids regulating TP‐isoprostane interaction. Our data revealed three key residues, i.e., Phe 184 , Phe 196 (unique for isoprostanes), and Asp 193 as important for 8‐iso‐PGF 2 α binding and functional activity. In addition, 8‐iso‐PGF 2 α was found to induce human platelet shape change through a TP‐mediated, RhoA kinase‐dependent mechanism. Interestingly, 8‐iso‐PGF 2 α also elevated platelet cAMP levels. This increase in cAMP was not mediated by either TP or FP (which normally do not couple to Gs), nor was this increase mediated by other prostanoid receptors which are known to couple to Gs, i.e., IP, DP or EP. In summary, these studies: 1. identify the isoprostane‐TP molecular coordination sites; 2. demonstrate that 8‐iso‐PGF 2 α can signal by TP‐dependent and TP‐independent mechanisms; and 3. suggest that the TP‐independent mechanism proceeds through a novel Gs‐coupled isoprostane receptor. This work was supported by a grant from the National Institutes of Health (G.C.L) and a predoctoral fellowship from the American Heart Association (F.T.K).