title: CoA-independent transacylase activity is increased in human neutrophils after treatment with tumor necrosis factor alpha.
abstract: CoA-independent transacylase (CoA-IT) appears to play a critical role in lipid mediator generation by rapidly moving arachidonate (AA) between phospholipid pools during cell activation. Tumor necrosis factor-alpha (TNF) pretreatment of human neutrophils increases agonist-induced production of inflammatory mediators. The current study tested if the TNF-induced increase in lipid mediator production may be, in part, due to altered CoA-IT activity. Neutrophils were treated with TNF (250 U/ml, 30 min), homogenates prepared, and CoA-IT activity measured by the ability of these homogenates to acylate 1-[3H]alkyl-2-lyso-sn-glycero-3-phosphocholine (GPC). There was an increased CoA-IT activity, from 9.1 +/- 1.1 to 13.7 +/- 1.4 pmol/mg per min in control vs. TNF-treated samples, respectively. Varying the concentration of 1-alkyl-2-lyso-GPC revealed an increased CoA-IT activity in microsomes that was due to an increased Vmax, from 26 to 54 pmol/mg per min. The ability of TNF to increase CoA-IT activity was concentration-dependent, with maximal response observed at 25 U/ml. This effect on CoA-IT appears to be specific, in that TNF treatment of neutrophils had no effect on CoA-dependent acylation of 1-acyl-2-lyso-sn-glycero-3-phosphocholine, using either AA-CoA or linolenoyl-CoA as substrates. In the intact cell, the movement of [3H]AA from other phospholipids into PE in fMLP-stimulated neutrophils was greatly enhanced after TNF treatment, demonstrating a functional consequence of increased CoA-IT activity. In addition, TNF treatment doubled platelet-activating factor production in response to the chemotactic peptide fMLP, as measured by [3H]acetate incorporation, while the response to A23187 remained unchanged. Taken together, these results provide the first evidence of modulation of CoA-IT activity by a proinflammatory cytokine and suggest that one mechanism for augmented lipid mediator formation is through increases in CoA-IT activity.
authors.affiliation: Department of Inflammation and Respiratory Pharmacology, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406.
authors.forename: J D
authors.initials: JD
authors.lastname: Winkler
authors.forename: C M
authors.initials: CM
authors.lastname: Sung
authors.forename: L
authors.initials: L
authors.lastname: Huang
authors.forename: F H
authors.initials: FH
authors.lastname: Chilton
mesh_terms: D000217:Acyltransferases / Q000378:metabolism*; D016718:Arachidonic Acid / Q000378:metabolism; D006801:Humans; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D010743:Phospholipids / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*; D015854:Up-Regulation
chemical_list: D010743:Phospholipids; D014409:Tumor Necrosis Factor-alpha; D016718:Arachidonic Acid; D009240:N-Formylmethionine Leucyl-Phenylalanine; D000217:Acyltransferases; C039766:arachidonyl transacylase