期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 280, 期 1, 页码 644-653出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M410229200
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资金
- NHLBI NIH HHS [HL57900, HL56595] Funding Source: Medline
- NIAID NIH HHS [AI38343] Funding Source: Medline
Studies with inhibitors have implicated protein kinase C (PKC) in the adhesive functions of integrin alpha(IIb)beta(3) in platelets, but the responsible PKC isoforms and mechanisms are unknown. alpha(IIb)beta(3) interacts directly with tyrosine kinases c-Src and Syk. Therefore, we asked whether alpha(IIb)beta(3) might also interact with PKC. Of the several PKC isoforms expressed in platelets, only PKCbeta co-immunoprecipitated with alpha(IIb)beta(3) in response to the interaction of platelets with soluble or immobilized fibrinogen. PKCbeta recruitment to alpha(IIb)beta(3) was accompanied by a 9-fold increase in PKC activity in alpha(IIb)beta(3) immunoprecipitates. RACK1, an intracellular adapter for activated PKCbeta, also co-immunoprecipitated with alphaIIbbeta3, but in this case, the interaction was constitutive. Broad spectrum PKC inhibitors blocked both PKCbeta recruitment to alpha(IIb)beta(3) and the spread of platelets on fibrinogen. Similarly, mouse platelets that are genetically deficient in PKCbeta spread poorly on fibrinogen, despite normal agonist-induced fibrinogen binding. In a Chinese hamster ovary cell model system, adhesion to fibrinogen caused green fluorescent protein-PKCbetaI to associate with alpha(IIb)beta(3) and to co-localize with it at lamellipodial edges. These responses, as well as Chinese hamster ovary cell migration on fibrinogen, were blocked by the deletion of the beta(3) cytoplasmic tail or by co-expression of a RACK1 mutant incapable of binding to beta(3). These studies demonstrate that the interaction of alpha(IIb)beta(3) with activated PKCbeta is regulated by integrin occupancy and can be mediated by RACK1 and that the interaction is required for platelet spreading triggered through alpha(IIb)beta(3). Furthermore, the studies extend the concept of alpha(IIb)beta(3) as a scaffold for multiple protein kinases that regulate the platelet actin cytoskeleton.
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