Phosphorylation of cPLA₂α at Ser⁵⁰⁵ Is Necessary for Its Translocation to PtdInsP₂-Enriched Membranes

Group IVA cytosolic phospholipase A₂α (cPLA₂α) is a key enzyme in physiology and pathophysiology because it constitutes a rate-limiting step in the pathway for the generation of pro- and anti-inflammatory eicosanoid lipid mediators. cPLA₂α activity is tightly regulated by multiple factors, including the intracellular Ca²⁺ concentration, phosphorylation reactions, and cellular phosphatidylinositol (4,5) bisphosphate levels (PtdInsP₂). In the present work, we demonstrate that phosphorylation of the enzyme at Ser⁵⁰⁵ is an important step for the translocation of the enzyme to PtdInsP₂-enriched membranes in human cells. Constructs of eGFP-cPLA₂ mutated in Ser⁵⁰⁵ to Ala (S505A) exhibit a delayed translocation in response to elevated intracellular Ca²⁺, and also in response to increases in intracellular PtdInsP₂ levels. Conversely, translocation of a phosphorylation mimic mutant (S505E) is fully observed in response to cellular increases in PtdInsP₂ levels. Collectively, these results suggest that phosphorylation of cPLA₂α at Ser⁵⁰⁵ is necessary for the enzyme to translocate to internal membranes and mobilize arachidonic acid for eicosanoid synthesis.