title: Beta-cell calcium-independent group VIA phospholipase A(2) (iPLA(2)beta): tracking iPLA(2)beta movements in response to stimulation with insulin secretagogues in INS-1 cells.
abstract: 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.
authors.affiliation: Department of Medicine, Washington University School of Medicine, St Louis, Missouri 63110, USA.
authors.forename: Shunzhong
authors.initials: S
authors.lastname: Bao
authors.forename: Chun
authors.initials: C
authors.lastname: Jin
authors.forename: Sheng
authors.initials: S
authors.lastname: Zhang
authors.forename: John
authors.initials: J
authors.lastname: Turk
authors.forename: Zhongmin
authors.initials: Z
authors.lastname: Ma
authors.forename: Sasanka
authors.initials: S
authors.lastname: Ramanadham
mesh_terms: D002118:Calcium / Q000502:physiology; D002460:Cell Line; D017931:DNA Primers; D004721:Endoplasmic Reticulum / Q000502:physiology; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D054522:Group VI Phospholipases A2; D006801:Humans; D007515:Islets of Langerhans / Q000201:enzymology; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D010741:Phospholipases A / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection
chemical_list: D017931:DNA Primers; D008164:Luminescent Proteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D010741:Phospholipases A; D054522:Group VI Phospholipases A2; C516475:PLA2G6 protein, human; D002118:Calcium