A β 1 integrin signaling pathway involving Src‐family kinases, Cbl and PI‐3 kinase is required for macrophage spreading and migration

We have used mutant macrophages which are deficient in expression of Src‐family kinases to define an integrin signaling pathway that is required for macrophage adhesion and migration. Following ligation of surface integrins by fibronectin, the p120 c‐cbl (Cbl) protein rapidly becomes tyrosine phosphorylated and associated with the Src‐family kinases Fgr and Lyn. In hck −/− fgr −/− lyn −/− triple mutant cells, which are defective in spreading on fibronectin‐coated surfaces in vitro and show impaired migration in vivo , Cbl tyrosine phosphorylation is blocked, Cbl protein levels are low, adhesion‐dependent translocation of Cbl to the membrane is impaired and Cbl‐associated, membrane‐localized phosphatidylinositol 3 (PI‐3)‐kinase activity is dramatically reduced. In contrast, adhesiondependent activation of total cellular PI‐3 kinase activity is normal in mutant cells, demonstrating that it is the membrane‐associated fraction of PI‐3 kinase which is most critical in regulating actin cytoskeletal rearrangements that lead to cell spreading. Treatment of wild‐type cells with the Src‐family‐specific inhibitor PP1, Cbl antisense oligonucleotides or pharmacological inhibitors of PI‐3 kinase blocks cell spreading on fibronectin surfaces. These data provide a molecular description for the role of Src‐family kinases Hck, Fgr and Lyn in β 1 ‐integrin signal transduction in macrophages.