DISSECTION OF PATHWAYS IMPLICATED IN INTEGRIN-MEDIATED ACTIN CYTOSKELETON ASSEMBLY - INVOLVEMENT OF PROTEIN-KINASE-C, RHO GTPASE, AND TYROSINE PHOSPHORYLATION

A panel of antibodies to the alpha IIb beta 3 integrin was used to pro mote adhesion of Chinese hamster ovary cells transfected with the alph a IIb beta 3 fibrinogen receptor. While some alpha IIb beta 3 antibodi es were not able to induce p125 focal adhesion kinase (p125FAK) tyrosi ne phosphorylation, all the antibodies equally support cell adhesion b ut not spreading and assembly of actin stress fibers, Absence of stres s fibers was also obtained by plating on antibodies directed to the ha mster beta 1 integrin. In contrast, cells plated on matrix proteins sp read organizing actin stress fibers. Treatment with phorbol esters pho rbol 12-myristate 13-acetate (PMA) induced cells to spread on antibodi es-coated dishes but not to organize actin in stress fibers, The combi nation of PMA and cytotoxic necrotizing factor 1 (CNF1), a specific Rh o activator, induced cell spreading and organization of stress fibers. PMA or the combination of PMA and CNF1 also increases tyrosine phosph orylation of p125FAK in response to antibodies that were otherwise una ble to trigger this response, These data show that: 1) matrix proteins and antibodies differ in their ability to induce integrin-dependent a ctin cytoskeleton organization (while matrix induced stress fibers for mation, antibodies did not); 2) p125FAK tyrosine phosphorylation is in sufficient per se to trigger actin stress fibers formation since antib odies that activate p125FAK tyrosine phosphorylation did not lead to a ctin stress fibers assembly; and 3) the inability of anti-integrin ant ibodies to trigger stress fibers organization is overcome by concomita nt activation of the protein kinase C (PKC) and Rho pathways; PKC acti vation leads to cell spreading and Rho activation is required to organ ize actin stress fibers.