Integrin-Independent Role of CalDAG-GEFI in Neutrophil Chemotaxis

Abstract

Chemotaxis and integrin activation are essential processes for neutrophil transmigration in response to injury. CalDAG-GEFI plays a key role in the activation of β1, β2, and β3 integrins in platelets and neutrophils by exchanging a GDP for a GTP on Rap1. Here, we explored the role of CalDAG-GEFI and Rap1b in integrin-independent neutrophil chemotaxis. In a transwell assay, (CalDAG-GEFI^{−/−}) neutrophils had a 46% reduction in transmigration compared with WT in response to a low concentration of (LTB_4). Visualization of migrating neutrophils in the presence of 10 mM EDTA revealed that (CalDAG-GEFI^{−/−}) neutrophils had abnormal chemotactic behavior compared with WT neutrophils, including reduced speed and directionality. Interestingly, (Rap1b^{−/−}) neutrophils had a similar phenotype in this assay, suggesting that CalDAG-GEFI may be acting through Rap1b. We investigated whether the deficit in integrin-independent chemotaxis in (CalDAG-GEFI^{−/−}) neutrophils could be explained by defective cytoskeleton rearrangement. Indeed, we found that CalDAG-GEFI−/− neutrophils had reduced formation of F-actin pseudopodia after (LTB_4) stimulation, suggesting that they have a defect in polarization. Overall, our studies show that CalDAG-GEFI helps regulate neutrophil chemotaxis, independent of its established role in integrin activation, through a mechanism that involves actin cytoskeleton and cellular polarization.