title: Multiple C-terminal serine phosphorylation accompanies both protein kinase C-dependent and -independent activation of cytosolic 85 kDa phospholipase A2 in macrophages.
abstract: Exposure of mouse macrophages to either phorbol ester or certain bacteria was previously shown to cause increased phosphorylation of the cytosolic 85 kDa phospholipase A2 as well as a stable increase in its catalytic activity. We have now attempted to map the major phosphorylation sites on the enzyme in such cells. Phosphorylation occurred on serine residues without a detectable increase in either phosphothreonine or phosphotyrosine. After CNBr cleavage five fragments showed increased 32P labelling. Among those the most heavily labelled fragment was identified as the most C-terminal (residues 698-749), containing six serine residues. This was true whether phorbol ester or bacteria, causing protein kinase C-independent phospholipase A2 activation, was used as stimulus. The heavy phosphorylation of the most C-terminal fragment and an analysis of tryptic peptides derived from it suggested that more than one of the six serine residues became phosphorylated. Smaller increases also occurred in other CNBr-cleaved fragments from the C-terminal part of the protein, including that carrying Ser-505, a known target of the mitogen-activated protein kinase ERK-2 (extracellular-signal regulated kinase). Dexamethasone treatment (1-100 nM for 20 h), which was earlier shown to dose-dependently down-regulate the 85 kDa phospholipase A2 and its activation by phorbol ester and zymosan, was here shown also to counteract the protein kinase C-independent activation and arachidonate release elicited by bacteria. It remains to be determined whether all phosphorylation sites are equally affected under those conditions.
authors.affiliation: Department of Cell and Molecular Biology, Lund University, P.O. Box 94, SE-221 00 Lund, Sweden.
authors.forename: J
authors.initials: J
authors.lastname: Wijkander
authors.forename: K
authors.initials: K
authors.lastname: Gewert
authors.forename: U
authors.initials: U
authors.lastname: Svensson
authors.forename: E
authors.initials: E
authors.lastname: Holst
authors.forename: R
authors.initials: R
authors.lastname: Sundler
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016718:Arachidonic Acid / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D005260:Female; D016967:Fusobacterium nucleatum / Q000502:physiology; D006194:Gardnerella vaginalis / Q000502:physiology; D017737:Macrophages, Peritoneal / Q000201:enzymology*; D051379:Mice; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000737:chemistry / Q000378:metabolism*; D054467:Phospholipases A2; D010748:Phosphopeptides / Q000032:analysis; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D012694:Serine / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D010748:Phosphopeptides; D016718:Arachidonic Acid; D012694:Serine; D003907:Dexamethasone; D011493:Protein Kinase C; D010741:Phospholipases A; D054467:Phospholipases A2; D013755:Tetradecanoylphorbol Acetate