Perinuclear localization of cytosolic phospholipase A₂α is important but not obligatory for coupling with cyclooxygenases

Abstract 

In response to Ca²⁺ signaling, cytosolic phospholipase A₂α (cPLA₂α) translocates from the cytosol to the perinuclear membrane, where downstream eicosanoid-synthetic enzymes, such as cyclooxygenase (COX), are localized. Although the spatiotemporal perinuclear colocalization of cPLA₂α and COXs has been proposed to be critical for their functional coupling leading to prostanoid production, definitive evidence for this paradigm has remained elusive. To circumstantiate this issue, we took advantage of a chimeric cPLA₂α mutant harboring the C2 domain of protein kinase Cα, which translocates to the plasma membrane following cell activation. Transfection analyses of the native or chimeric cPLA₂α in combination with COX-1 or COX-2 revealed that, even though the arachidonate-releasing capacities of native and mutant cPLA₂α were comparable, prostaglandin production by mutant cPLA₂α was markedly impaired as compared with that by native cPLA₂α. We thus conclude that the perinuclear localization of cPLA₂α is preferential, even if not obligatory, for efficient coupling with COXs.

Keywords:  Phospholipase A2, Cyclooxygenase, Arachidonic acid, Prostaglandin, C2 domain

Abbreviations:  PLA₂, phospholipase A₂, cPLA₂, cytosolic phospholipase A₂, sPLA₂, secretory PLA₂, iPLA₂, Ca²⁺-independent PLA₂, COX, cyclooxygenase, 5-LO, 5-lipoxygenase, AA, arachidonic acid, PG, prostaglandin, LT, leukotriene, PKC, protein kinase C, EGFP, enhanced green fluorescent protein, IL-1, interleukin-1, FCS, fetal calf serum, WT, wild-type, HEK, human embryonic kidney, PC, phosphatidylcholine