Rap1 small GTPase is essential for maintaining pulmonary endothelial barrier function in mice

Vascular permeability is dynamically but tightly controlled by vascular endothelial (VE)-cadherin-mediated endothelial cell–cell junctions to maintain homeostasis. Thus, impairments of VE-cadherin-mediated cell adhesions lead to hyperpermeability, promoting the development and progression of various disease processes. Notably, the lungs are a highly vulnerable organ wherein pulmonary inflammation and infection result in vascular leakage. Herein, we showed that Rap1, a small GTPase, plays an essential role for maintaining pulmonary endothelial barrier function in mice. Endothelial cell-specific Rap1a/Rap1b double knockout mice exhibited severe pulmonary edema. They also showed vascular leakage in the hearts, but not in the brains. En face analyses of the pulmonary arteries and 3D-immunofluorescence analyses of the lungs revealed that Rap1 potentiates VE-cadherin-mediated endothelial cell–cell junctions through dynamic actin cytoskeleton reorganization. Rap1 inhibits formation of cytoplasmic actin bundles perpendicularly binding VE-cadherin adhesions through inhibition of a Rho-ROCK pathway-induced activation of cytoplasmic nonmuscle myosin II (NM-II). Simultaneously, Rap1 induces junctional NM-II activation to create circumferential actin bundles, which anchor and stabilize VE-cadherin at cell–cell junctions. We also showed that the mice carrying only one allele of either Rap1a or Rap1b out of the two Rap1 genes are more vulnerable to lipopolysaccharide (LPS)-induced pulmonary vascular leakage than wild-type mice, while activation of Rap1 by administration of 007, an activator for Epac, attenuates LPS-induced increase in pulmonary endothelial permeability in wild-type mice. Thus, we demonstrate that Rap1 plays an essential role for maintaining pulmonary endothelial barrier functions under physiological conditions and provides protection against inflammation-induced pulmonary vascular leakage.