title: Rap-afadin axis in control of Rho signaling and endothelial barrier recovery.
abstract: Activation of the Rho GTPase pathway determines endothelial cell (EC) hyperpermeability after injurious stimuli. To date, feedback mechanisms of Rho down-regulation critical for barrier restoration remain poorly understood. We tested a hypothesis that Rho down-regulation and barrier recovery of agonist-stimulated ECs is mediated by the Ras family GTPase Rap1. Thrombin-induced EC permeability driven by rapid activation of the Rho GTPase pathway was followed by Src kinase-dependent phosphorylation of the Rap1-specific guanine nucleotide exchange factor (GEF) C3G, activation of Rap1, and initiation of EC barrier recovery. Knockdown experiments showed that Rap1 activation was essential for down-regulation of Rho signaling and actin stress fiber dissolution. Rap1 activation also enhanced interaction between adherens junction (AJ) proteins VE-cadherin and p120-catenin and stimulated AJ reannealing mediated by the Rap1 effector afadin. This mechanism also included Rap1-dependent membrane translocation of the Rac1-specific GEF Tiam1 and activation of Rac1-dependent peripheral cytoskeletal dynamics, leading to resealing of intercellular gaps. These data demonstrate that activation of the Rap1-afadin axis is a physiological mechanism driving restoration of barrier integrity in agonist-stimulated EC monolayers via negative-feedback regulation of Rho signaling, stimulation of actin peripheral dynamics, and reestablishment of cell-cell adhesive complexes.
authors.affiliation: Department of Medicine, Section of Pulmonary and Critical Medicine, Lung Injury Center, University of Chicago, Chicago, IL 60637, USA.
authors.forename: Anna A
authors.initials: AA
authors.lastname: Birukova
authors.forename: Xinyong
authors.initials: X
authors.lastname: Tian
authors.forename: Yufeng
authors.initials: Y
authors.lastname: Tian
authors.forename: Katherine
authors.initials: K
authors.lastname: Higginbotham
authors.forename: Konstantin G
authors.initials: KG
authors.lastname: Birukov
mesh_terms: D003599:Cytoskeleton / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008840:Microfilament Proteins / Q000378:metabolism*; D010766:Phosphorylation; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D013917:Thrombin / Q000494:pharmacology*; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism*; D020741:rho GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D008840:Microfilament Proteins; C108835:afadin; D013917:Thrombin; D020827:rap1 GTP-Binding Proteins; D020741:rho GTP-Binding Proteins