Rap1a activation by CalDAG‐GEFI and p38 MAPK is involved in E‐selectin‐dependent slow leukocyte rolling

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Abstract
Rolling leukocytes are exposed to different adhesion molecules and chemokines. Neutrophils rolling on E‐selectin induce integrin αLβ2‐mediated slow rolling on ICAM‐1 by activating a phospholipase C (PLC)γ2‐dependent and a separate PI3Kγ‐dependent pathway. E‐selectin‐signaling cooperates with chemokine signaling to recruit neutrophils into inflamed tissues. However, the distal signaling pathway linking PLCγ2 (Plcg2) to αLβ2‐activation is unknown. To identify this pathway, we used different Tat‐fusion‐mutants and gene‐deficient mice in intravital microscopy, autoperfused flow chamber, peritonitis, and biochemical studies. We found that the small GTPase Rap1 is activated following E‐selectin engagement and that blocking Rap1a in Pik3cg−/− mice by a dominant‐negative Tat‐fusion mutant completely abolished E‐selectin‐mediated slow rolling. We identified CalDAG‐GEFI (Rasgrp2) and p38 MAPK as key signaling intermediates between PLCγ2 and Rap1a. Gαi‐independent leukocyte adhesion to and transmigration through endothelial cells in inflamed postcapillary venules of the cremaster muscle were completely abolished in Rasgrp2−/− mice. The physiological importance of CalDAG‐GEFI in E‐selectin‐dependent integrin activation is shown by complete inhibition of neutrophil recruitment into the inflamed peritoneal cavity of Rasgrp2−/− leukocytes treated with pertussis toxin to block Gαi‐signaling. Our data demonstrate that Rap1a activation by p38 MAPK and CalDAG‐GEFI is involved in E‐selectin‐dependent slow rolling and leukocyte recruitment.