The role of protein disulfide isomerase in the post-ligation phase of β3 integrin-dependent cell adhesion

Abstract Introduction Protein disulfide isomerase (PDI) catalyzes disulfide bond exchange. It is crucial for integrin-mediated platelet adhesion and aggregation and disulfide bond exchange is necessary for α IIb β 3 and α v β 3 activation. However, the role of disulfide bond exchange and PDI in the post-ligation phase of α IIb β 3 and α v β 3 mediated cell adhesion has yet to be determined. Methods To investigate a possible such role, we expressed wild type (WT) human α IIb and either WT human β 3 , or β 3 harboring single or double cysteine to serine substitutions disrupting Cys473–Cys503 or Cys523–Cys544 bonds, in baby hamster kidney (BHK) cells, leading to expression of both human α IIb β 3 and a chimeric hamster/human α v β 3 . Adhesion to fibrinogen-coated wells was studied in the presence or absence of bacitracin, a PDI inhibitor, with and without an α v β 3 blocker. Results Flow cytometry showed WT and mutant α IIb β 3 expression in BHK cells and indicated that mutated α IIb β 3 receptors were constitutively active while WT α IIb β 3 was inactive. Both α IIb β 3 and α v β 3 integrins, WT and mutants, mediated adhesion to fibrinogen as shown by reduced but still substantial adhesion following treatment with the α v β 3 blocker. Mutated α IIb β 3 integrins disrupted in the Cys523–Cys544 bond still depended on PDI for adhesion as shown by the inhibitory effect of bacitracin in the presence of the α v β 3 blocker. Mutated integrins disrupted in the Cys473–Cys503 bond showed a similar trend. Conclusions PDI-mediated disulfide bond exchange plays a pivotal role in the post-ligation phase of α IIb β 3 -mediated adhesion to fibrinogen, while this step in α v β 3 -mediated adhesion is independent of disulfide exchange.