title: Tyrosine phosphorylation of phospholipase C-gamma 2 upon cross-linking of membrane Ig on murine B lymphocytes.
abstract: When membrane Ig (mIg) on the surface of B lymphocytes is cross-linked using anti-Ig antibodies, the enzyme phospholipase C (PLC) is activated to cleave inositol phospholipids. Tyrosine kinase inhibitors have been reported to inhibit this event. Therefore, we investigated the effect of cross-linking of mIg on the state of tyrosine phosphorylation of PLC activity in two murine B cell lines and in normal resting mouse B cells. Proteins from lysates of stimulated or unstimulated cells were immunoprecipitated with an antiphosphotyrosine antibody and subsequently assayed for PLC activity. Treatment of the B cell line WEHI-231 with anti-IgM led within 15 to 30 s to a 10- to 20-fold increase in tyrosine-phosphorylated PLC activity. Inositol trisphosphate generation by WEHI-231 cells stimulated under the same conditions demonstrated similar kinetics. Normal resting B cells treated with anti-IgM or anti-IgD demonstrated 2.5- and 4-fold increases, respectively, of tyrosine-phosphorylated PLC activity. To identify the isozyme of PLC that was phosphorylated, we immunoprecipitated PLC-gamma 1 or PLC-gamma 2 with specific antibodies and assessed the amount of tyrosine phosphorylation of these proteins by antiphosphotyrosine immunoblotting. Treatment of WEHI-231 or Bal17 cells with anti-IgM induced an increase in PLC-gamma 2 tyrosine phosphorylation over background levels. There was no detectable tyrosine phosphorylation of PLC-gamma 1 in treated or untreated WEHI-231 cells, whereas anti-IgM-treated Bal17 cells did exhibit low but detectable levels of tyrosine phosphorylation of PLC-gamma 1. In normal resting mouse B cells, there was no detectable PLC-gamma 1, but PLC-gamma 2 was abundant. These observations suggest that PLC-gamma 2 is a significant substrate for the mIg-activated protein tyrosine kinase and may be responsible for mediating mIg stimulation of inositol phospholipid hydrolysis in murine B cells.
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco 94143.
authors.forename: W M
authors.initials: WM
authors.lastname: Hempel
authors.forename: R C
authors.initials: RC
authors.lastname: Schatzman
authors.forename: A L
authors.initials: AL
authors.lastname: DeFranco
mesh_terms: D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D001402:B-Lymphocytes / Q000201:enzymology*; D019204:GTP-Binding Proteins / Q000502:physiology; D007075:Immunoglobulin M / Q000276:immunology / Q000502:physiology*; D007527:Isoenzymes / Q000032:analysis; D051379:Mice; D008815:Mice, Inbred Strains; D010766:Phosphorylation; D011947:Receptors, Antigen, B-Cell / Q000502:physiology*; D010738:Type C Phospholipases / Q000032:analysis / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism*
chemical_list: D000888:Antibodies, Anti-Idiotypic; D007075:Immunoglobulin M; D007527:Isoenzymes; D011947:Receptors, Antigen, B-Cell; D014443:Tyrosine; D010738:Type C Phospholipases; D019204:GTP-Binding Proteins