title: Protein phosphatase 2A carboxymethylation and regulatory B subunits differentially regulate mast cell degranulation.
abstract: Asthma is characterised by antigen-mediated mast cell degranulation resulting in secretion of inflammatory mediators. Protein phosphatase 2A (PP2A) is a serine/threonine protein phosphatase composed of a catalytic (PP2A-C) subunit together with a core scaffold (PP2A-A) subunit and a variable, regulatory (PP2A-B) subunit. Previous studies utilising pharmacological inhibition of protein phosphatases have suggested a positive regulatory role for PP2A in mast cell degranulation. In support of this we find that a high okadaic acid concentration (1μM) inhibits mast cell degranulation. Strikingly, we now show that a low concentration of okadaic acid (0.1μM) has the opposite effect, resulting in enhanced degranulation. Selective downregulation of the PP2A-Cα subunit by short hairpin RNA also enhanced degranulation of RBL-2H3 mast cells, suggesting that the primary role of PP2A is to negatively regulate degranulation. PP2A-B subunits are responsible for substrate specificity, and carboxymethylation of the PP2A-C subunit alters B subunit binding. We show here that carboxymethylation of PP2A-C is dynamically altered during degranulation and inhibition of methylation decreases degranulation. Moreover downregulation of the PP2A-Bα subunit resulted in decreased MK2 phosphorylation and degranulation, whilst downregulation of the PP2A-B'δ subunit enhanced p38 MAPK phosphorylation and degranulation. Taken together these data show that PP2A is both a positive and negative regulator of mast cell degranulation, and this differential role is regulated by carboxymethylation and specific PP2A-B subunit binding.
authors.affiliation: School of Biomedical Sciences, Faculty of Health, University of Newcastle, Callaghan, NSW 2308, Australia.
authors.forename: Gregory
authors.initials: G
authors.lastname: Kranias
authors.forename: Lauren F
authors.initials: LF
authors.lastname: Watt
authors.forename: Helen
authors.initials: H
authors.lastname: Carpenter
authors.forename: Jeff
authors.initials: J
authors.lastname: Holst
authors.forename: Russell
authors.initials: R
authors.lastname: Ludowyke
authors.forename: Stefan
authors.initials: S
authors.lastname: Strack
authors.forename: Alistair T R
authors.initials: AT
authors.lastname: Sim
authors.forename: Nicole M
authors.initials: NM
authors.lastname: Verrills
mesh_terms: D000818:Animals; D015550:Cell Degranulation / Q000502:physiology*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects; D008407:Mast Cells / Q000502:physiology*; D008745:Methylation; D008954:Models, Biological; D019319:Okadaic Acid / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D010766:Phosphorylation; D054648:Protein Phosphatase 2 / Q000378:metabolism*; D021122:Protein Subunits / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism
chemical_list: D021122:Protein Subunits; D034741:RNA, Small Interfering; D019319:Okadaic Acid; D048051:p38 Mitogen-Activated Protein Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2