Phosphorylation‐ and stimulus‐dependent inhibition of cellular 5‐lipoxygenase activity by nonredox‐type inhibitors

TLDR

Nonredox-type 5‑LO inhibitors such as ZM230487 and L‑739.010 potently suppress leukotriene biosynthesis even at low cellular peroxide levels. Compared with Ca²⁺‑mediated activation, 5‑LO phosphorylation pathways render the enzyme 10– to 100‑fold less sensitive to nonredox inhibitors ZM230487 and L‑739.010, whereas iron‑ligand and novel inhibitors remain equally effective, indicating that phosphorylation events specifically modulate inhibitor susceptibility.

Abstract

Nonredox-type 5-lipoxygenase (5-LO) inhibitors such as ZM230487 or L-739.010 potently suppress leukotriene biosynthesis at low cellular peroxide tone. Upper. Thus, compared with 5-LO product synthesis induced by the Ca2+-mobilizing agent ionophore A23187, cell stress-induced 5-LO product formation involving 5-LO kinase pathways required ~10- to 100-fold higher concentrations of ZM230487 or L-739.010 for comparable 5-LO inhibition. No such differences were observed for the iron ligand-type 5-LO inhibitor BWA4C or the novel-type 5-LO inhibitors hyperforin and 3-O-acetyl-11-keto-boswellic acid. Experiments using purified 5-LO revealed that Ca2+ is no prerequisite for potent enzyme inhibition by ZM230487, and exposure of PMNL to the combination of ionophore and cell stress did not restore potent 5-LO suppression. Intriguingly, a significant difference in the potency of nonredox-type inhibitors (but not of BWA4C) was determined between wild-type 5-LO and the mutant S271A/S663A-5-LO (lacking phosphorylation sites for ERK1/2 and MAPKAPK-2) in HeLa cells. Collectively, our data suggest that compared with Ca2+-mediated 5-LO product formation, enzyme activation involving 5-LO phosphorylation events specifically and strongly alters the susceptibility of 5-LO toward nonredox-type inhibitors in intact cells.