Evidence that group VIA cytosolic
calcium-independent
phospholipase A(2) (
iPLA(2)beta) participates in beta-cell signal transduction includes the observations that inhibition of
iPLA(2)beta with the
bromoenol lactone suicide substrate suppresses
glucose-stimulated insulin secretion and that overexpression of
iPLA(2)beta amplifies
insulin secretory responses in INS-1
insulinoma cells. Immunofluorescence analyses also reveal that
iPLA(2)beta accumulates in the perinuclear region of INS-1 cells stimulated with
glucose and
forskolin. To characterize this phenomenon further,
iPLA(2)beta was expressed as a fusion
protein with
enhanced green fluorescent protein (EGFP) in INS-1 cells so that movements of
iPLA(2)beta are reflected by changes in the subcellular distribution of green fluorescence. Stimulation of INS-1 cells overexpressing iPLA(2)beta-EGFP induced greater insulin secretion and punctate accumulation of iPLA(2)beta-EGFP fluorescence in the perinuclear region. To determine the identity of organelles with which
iPLA(2)beta might associate, colocalization of green fluorescence with fluorophores associated with specific trackers targeted to different subcellular organelles was examined. Such analyses reveal association of iPLA(2)beta-EGFP fluorescence with the ER and Golgi compartments. Arachidonate-containing
plasmenylethanolamine phospholipid species are abundant in beta-cell endoplasmic reticulum (ER) and are excellent substrates for
iPLA(2)beta.
Arachidonic acid produced by iPLA(2)beta-catalyzed hydrolysis of their substrates induces release of Ca(2+) from ER stores-an event thought to participate in
glucose-stimulated insulin secretion.