Receptors for the 5-oxo class of eicosanoids in neutrophils.
5-Hydroxy- and 5-oxo-eicosatetraenoate (5-HETE and 5-oxoETE) activate polymorphonuclear neutrophils (PMNs) through a common, receptor-like recognition system. To define this system, we examined the interaction of these eicosanoids with human PMNs. PMNs esterified 5-[3H]HETE to glycerolipids at 37 and 4 degreesC. At 37 but not 4 degreesC, the cells also hydroxylated the label to 5, 20-[3H]diHETE. The acyl:CoA synthetase blocker, triacsin C, inhibited esterification but also led to an increase in the hydroxylation of the label. PMNs processed 5-[3H]oxoETE through the same pathways but only or principally after reducing it to 5-[3H]HETE (37 or 4 degreesC). In the presence of these varying metabolic reactions, PMNs (37 or 4 degreesC; +/- triacsin C) could not be shown to receptor bind either radiolabel. Plasma membranes isolated from PMNs esterified but unlike whole cells did not reduce or hydroxylate 5-[3H]oxoETE. Triacsin C blocked esterification, thereby rendering the membranes unable to metabolize this radiolabel. Indeed, triacsin C-treated membranes bound (Kd = 3.8 nM) 5-[3H]oxoETE specifically and reversibly to 86 pmol of sites per 25 micrograms of membrane protein. 5-OxoETE, 5-HETE, and 5,15-diHETE displaced this binding at concentrations correlating with their potency in eliciting PMN Ca2+ transients. GTP and GTPgammaS, but not ATP or ATPgammaS, also reduced 5-[3H]oxoETE binding, whereas 15-HETE, leukotriene B4, platelet-activating factor, IL-8, C5a, and N-formyl-Met-Leu-Phe lacked this effect. We conclude that PMNs and their plasma membranes use an acyl:CoA synthetase-dependent route to esterify 5-HETE and 5-oxoETE into lipids. Blockade of the synthetase uncovers cryptic plasmalemma sites that bind 5-oxoETE with exquisite specificity. These sites apparently mediate responses to the 5-oxo class of eicosanoids and are likely members of the serpentine superfamily of G protein-linked receptors.
O'Flaherty
J T
JT
Departments of Internal Medicine, Wake Forest University Medical Center, Winston-Salem, North Carolina 27157, USA. joflaher@wfubmc.edu
Taylor
J S
JS
Thomas
M J
MJ
D002462:Cell Membrane / Q000378:metabolism; D015777:Eicosanoids / Q000378:metabolism*; D006801:Humans; D009504:Neutrophils / Q000378:metabolism*; D011485:Protein Binding; D018055:Receptors, Eicosanoid / Q000378:metabolism*; D014226:Triazenes / Q000494:pharmacology; D014316:Tritium
D015777:Eicosanoids; D018055:Receptors, Eicosanoid; D014226:Triazenes; D014316:Tritium; C034613:triacsin C