title: Inhibition of antigen and calcium ionophore induced secretion from RBL-2H3 cells by phosphatase inhibitors.
abstract: The role of serine/threonine protein phosphatases PP1 and PP2A in mast cell secretion was investigated using the phosphatase inhibitors okadaic acid and calyculin A. Calyculin A (5-25 nm) inhibited antigen-induced secretion from a rat mucosal mast cell line (RBL-2H3) when added in conjunction with the activator. Okadaic acid (250-1000 nm) inhibited secretion only when added before activation and did so in a time- and concentration-dependent manner. Both inhibitors caused the cells to become rounder, but only calyculin A induced membrane blebbing and a loss of adherence. Okadaic acid also inhibited secretion induced by the calcium ionophore A23187, in the presence or absence of PMA, indicating that the phosphatase inhibitors act on a component of the secretory pathway downstream of calcium mobilization. Okadaic acid increased the phosphorylation of a number of proteins, as did an analogue methyl okadaate, which also inhibited secretion, but less effectively. Okadaic acid induced the phosphorylation of triton-insoluble proteins of 55, 18 and 16 kDa. The 55 kDa protein was identified as vimentin and okadaic acid induced its partial translocation to the triton-soluble fraction. Our data indicate that full secretory function in mucosal mast cells requires phosphatase activity.
authors.affiliation: Centre for Immunology, St Vincent's Hospital, University of New South Wales, Sydney, Australia.
authors.forename: R I
authors.initials: RI
authors.lastname: Ludowyke
authors.forename: K
authors.initials: K
authors.lastname: Warton
authors.forename: L L
authors.initials: LL
authors.lastname: Scurr
mesh_terms: D000818:Animals; D000941:Antigens / Q000276:immunology*; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D048429:Cell Size / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D008387:Marine Toxins; D008407:Mast Cells / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism*; D019319:Okadaic Acid / Q000494:pharmacology; D010080:Oxazoles / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014746:Vimentin / Q000187:drug effects / Q000378:metabolism
chemical_list: D000941:Antigens; D004791:Enzyme Inhibitors; D007476:Ionophores; D008387:Marine Toxins; D010080:Oxazoles; D014746:Vimentin; D019319:Okadaic Acid; D000001:Calcimycin; C059041:calyculin A; D010749:Phosphoprotein Phosphatases; D002118:Calcium