title: Activation of SRC family kinases in human neutrophils. Evidence that p58C-FGR and p53/56LYN redistributed to a Triton X-100-insoluble cytoskeletal fraction, also enriched in the caveolar protein caveolin, display an enhanced kinase activity.
abstract: Protein tyrosine phosphorylation is one of the signals involved in stimulation of neutrophil (PMN) functions. We found that phorbol myristate acetate (PMA) activates the src family tyrosine kinases p58c-fgr and p53/56lyn in suspended PMNs. Moreover, we found that up to about 20% of p58c-fgr and p53/56lyn redistribute to a Triton X-100-insoluble fraction after PMA stimulation, and it is this fraction of the two kinases which diplays an increased activity. These changes of p58c-fgr and p53/56lyn distribution and activity correlate with tyrosine phosphorylation of endogenous substrates. In fact, in PMA-stimulated PMNs tyrosine phosphorylated proteins are mostly recovered in a Triton-insoluble cell fraction. To separate cytoskeletal from caveolar structures, which both display Triton X-100-insolubility, we used the detergent n-octyl beta-D-glucopyranoside (OGP) which solubilises components of caveolae. We found that the caveolae marker protein, caveolin, as well as the cytoskeletal protein alph-actinin and p58c-fgr and p53/56lyn, is insoluble in OGP. These findings suggest that PMA stimulation promotes the formation of multimolecular complexes containing cytoskeletal proteins, caveolin-containing structures and src family protein tyrosine kinases. Moreover, they show that p58c-fgr and p53/56lyn associated with this multimolecular complex display an enhanced kinase activity.
authors.affiliation: Institute of General Pathology, University of Verona, Italy.
authors.forename: S R
authors.initials: SR
authors.lastname: Yan
authors.forename: L
authors.initials: L
authors.lastname: Fumagalli
authors.forename: G
authors.initials: G
authors.lastname: Berton
mesh_terms: D000185:Actinin / Q000032:analysis; D051242:Caveolin 1; D022461:Caveolins; D002458:Cell Fractionation; D003599:Cytoskeleton / Q000737:chemistry* / Q000201:enzymology; D003902:Detergents; D004789:Enzyme Activation / Q000187:drug effects; D005960:Glucosides; D006801:Humans; D008565:Membrane Proteins / Q000032:analysis*; D018375:Neutrophil Activation / Q000187:drug effects; D009504:Neutrophils / Q000737:chemistry / Q000201:enzymology*; D017830:Octoxynol; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000032:analysis / Q000378:metabolism*; D012995:Solubility; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000032:analysis / Q000378:metabolism*
chemical_list: C495540:CAV1 protein, human; D051242:Caveolin 1; D022461:Caveolins; D003902:Detergents; D005960:Glucosides; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D000185:Actinin; C018619:octyl-beta-D-glucoside; D014443:Tyrosine; D017830:Octoxynol; C060902:lyn protein-tyrosine kinase; C053102:proto-oncogene proteins c-fgr; D019061:src-Family Kinases; D013755:Tetradecanoylphorbol Acetate