Open AccessA functional siRNA screen identifies RhoGTPase-associated genes involved in thrombin-induced endothelial permeability
Author(s) -
Joana Amado-Azevedo,
Renée X. de Menezes,
Geerten P. van Nieuw Amerongen,
Victor W.M. van Hinsbergh,
Peter L. Hordijk
Publication year - 2018
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0201231
Subject(s) - adherens junction , microbiology and biotechnology , thrombin , gene silencing , small interfering rna , vascular permeability , endothelial stem cell , biology , effector , actin cytoskeleton , endothelium , cytoskeleton , immunology , gene , cell , cadherin , transfection , biochemistry , platelet , genetics , in vitro , endocrinology
Thrombin and other inflammatory mediators may induce vascular permeability through the disruption of adherens junctions between adjacent endothelial cells. If uncontrolled, hyperpermeability leads to an impaired barrier, fluid leakage and edema, which can contribute to multi-organ failure and death. RhoGTPases control cytoskeletal dynamics, adhesion and migration and are known regulators of endothelial integrity. Knowledge of the precise role of each RhoGTPase, and their associated regulatory and effector genes, in endothelial integrity is incomplete. Using a combination of a RNAi screen with electrical impedance measurements, we quantified the effect of individually silencing 270 Rho-associated genes on the barrier function of thrombin-activated, primary endothelial cells. Known and novel RhoGTPase-associated regulators that modulate the response to thrombin were identified (RTKN, TIAM2, MLC1, ARPC1B, SEPT2, SLC9A3R1, RACGAP1, RAPGEF2, RHOD, PREX1, ARHGEF7, PLXNB2, ARHGAP45, SRGAP2, ARHGEF5). In conclusion, with this siRNA screen, we confirmed the roles of known regulators of endothelial integrity but also identified new, potential key players in thrombin-induced endothelial signaling.