title: Alteration in the activation state of new inflammation-associated targets by phospholipase A2-activating protein (PLAA).
abstract: Phospholipase A(2) (PLA(2))-activating protein (PLAA) is a novel signaling molecule that regulates the production of prostaglandins (PGE(2)) and tumor necrosis factor (TNF)-alpha. To characterize the function of native PLAA in situ, we generated HeLa (Tet-off) cells overexpressing plaa (plaa(high)) and control (plaa(low)) cells, with the plaa gene in opposite orientation in the latter construct. The plaa(high) cells produced significantly more PGE(2) and interleukin (IL)-6 compared to plaa(low) cells in response to TNF-alpha. There was an increased activation and/or expression of cytosolic PLA(2), cyclooxgenase-2, and NF-kappaB after induction of plaa(high) cells with TNF-alpha compared to the respective plaa(low) cells. Microarray analysis of plaa(high) cells followed by functional assays revealed increased production of proinflammatory cytokine IL-32 and a decrease in the production of annexin A4 and clusterin compared to plaa(low) cells. We demonstrated the role of annexin A4 as an inhibitor of PLA(2) and showed that addition of exogeneous clusterin limited the production of PGE(2) from plaa(high) cells. To understand regulation of plaa gene expression, we used a luciferase reporter system in HeLa cells and identified one stimulatory element, with Sp1 binding sites, and one inhibitory element, in exon 1 of the plaa gene. By using decoy DNA oligonucleotides to Sp1 and competitive binding assays, we showed that Sp1 maintains basal expression of the plaa gene and binds to the above-mentioned stimulatory element. We demonstrated for the first time that the induction of native PLAA by TNF-alpha can perpetuate inflammation by enhancing activation of PLA(2) and NF-kappaB.
authors.affiliation: Department of Microbiology & Immunology, The University of Texas Medical Branch, Galveston, TX 77555-1070, United States.
authors.forename: Fan
authors.initials: F
authors.lastname: Zhang
authors.forename: Jian
authors.initials: J
authors.lastname: Sha
authors.forename: Thomas G
authors.initials: TG
authors.lastname: Wood
authors.forename: Cristi L
authors.initials: CL
authors.lastname: Galindo
authors.forename: Harold R
authors.initials: HR
authors.lastname: Garner
authors.forename: Mark F
authors.initials: MF
authors.lastname: Burkart
authors.forename: Giovanni
authors.initials: G
authors.lastname: Suarez
authors.forename: Johanna C
authors.initials: JC
authors.lastname: Sierra
authors.forename: Stacy L
authors.initials: SL
authors.lastname: Agar
authors.forename: Johnny W
authors.initials: JW
authors.lastname: Peterson
authors.forename: Ashok K
authors.initials: AK
authors.lastname: Chopra
mesh_terms: D017314:Annexin A4 / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D051152:Clusterin / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D004789:Enzyme Activation; D015870:Gene Expression; D020869:Gene Expression Profiling; D006367:HeLa Cells; D006801:Humans; D007249:Inflammation / Q000209:etiology / Q000235:genetics / Q000378:metabolism*; D015850:Interleukin-6 / Q000096:biosynthesis; D007378:Interleukins / Q000235:genetics / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D054467:Phospholipases A2 / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction; D016329:Sp1 Transcription Factor / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
chemical_list: D017314:Annexin A4; C495242:CLU protein, human; D051152:Clusterin; D017931:DNA Primers; C496971:IL32 protein, human; C508600:IL6 protein, human; D015850:Interleukin-6; D007378:Interleukins; D016328:NF-kappa B; D011506:Proteins; D012333:RNA, Messenger; D016329:Sp1 Transcription Factor; D014409:Tumor Necrosis Factor-alpha; C055324:phospholipase A2-activating protein; D054467:Phospholipases A2