ICAM-2 regulates vascular permeability and N-cadherin localization through ezrin-radixin-moesin (ERM) proteins and Rac-1 signalling

Abstract

BackgroundEndothelial junctions control functions such as permeability, angiogenesis and contact inhibition. VE-Cadherin (VECad) is essential for the maintenance of intercellular contacts. In confluent endothelial monolayers, N-Cadherin (NCad) is mostly expressed on the apical and basal membrane, but in the absence of VECad it localizes at junctions. Both cadherins are required for vascular development. The intercellular adhesion molecule (ICAM)-2, also localized at endothelial junctions, is involved in leukocyte recruitment and angiogenesis.ResultsIn human umbilical vein endothelial cells (HUVEC), both VECad and NCad were found at nascent cell contacts of sub-confluent monolayers, but only VECad localized at the mature junctions of confluent monolayers. Inhibition of ICAM-2 expression by siRNA caused the appearance of small gaps at the junctions and a decrease in NCad junctional staining in sub-confluent monolayers. Endothelioma lines derived from WT or ICAM-2-deficient mice (IC2neg) lacked VECad and failed to form junctions, with loss of contact inhibition. Re-expression of full-length ICAM-2 (IC2 FL) in IC2neg cells restored contact inhibition through recruitment of NCad at the junctions. Mutant ICAM-2 lacking the binding site for ERM proteins (IC2 ΔERM) or the cytoplasmic tail (IC2 ΔTAIL) failed to restore junctions. ICAM-2-dependent Rac-1 activation was also decreased in these mutant cell lines. Barrier function, measured in vitro via transendothelial electrical resistance, was decreased in IC2neg cells, both in resting conditions and after thrombin stimulation. This was dependent on ICAM-2 signalling to the small GTPase Rac-1, since transendothelial electrical resistance of IC2neg cells was restored by constitutively active Rac-1. In vivo, thrombin-induced extravasation of FITC-labeled albumin measured by intravital fluorescence microscopy in the mouse cremaster muscle showed that permeability was increased in ICAM-2-deficient mice compared to controls.ConclusionsThese results indicate that ICAM-2 regulates endothelial barrier function and permeability through a pathway involving N-Cadherin, ERMs and Rac-1.

A functional siRNA screen identifies RhoGTPase-associated genes involved in thrombin-induced endothelial permeability

“…Using a RNAi approach in conjunction with electrical impedance measurements of the endothelial barrier, we analyzed the impact of silencing 270 genes (RhoGTPAses, RhoGEFs, RhoGAPs, GDIs, Effectors and Rho-Associated genes) on the response to thrombin. Our results confirm known regulators such as the RhoA-GEFs LARG [ 24 ] and GEF-H1 [ 36 ], the Gα protein GNA12 [ 37 ], the ROCK substrate Moesin [ 38 , 39 ], and the GTPase RND3 [ 40 ]. Interestingly, our analysis also uncovers interesting, potentially novel regulators of the endothelial barrier including the GTPase RhoD, and the GEFs TIAM2, ArhGEF5, ArhGEF7 and PLXNB3.…”

Tumor necrosis factor-α requires Ezrin to regulate the cytoskeleton and cause pulmonary microvascular endothelial barrier damage

“…Secondly, the specific mechanism of p-Ezrin in regulating the increase of cell permeability induced by TNF-α is still not fully clear, especially the mechanism of the Ezrin-related signal transduction pathway. Also, potential cooperation between related signal transduction pathways needs to be further investigated, especially the crosstalk effect of ICAM-2 and Rac1-Ezrin signaling ( Amsellem et al, 2014 ).…”

Proteomic analysis of physiological versus pathological cardiac remodeling in animal models expressing mutations in myosin essential light chains

“…Changes in the levels of important cytoskeletal proteins included cofilin-2, meosin, and cytoplasmic dynein 1 heavy chain 1 (Table 5). Moesin is an important cytoskeletal protein which has been shown to interact with ezrin and radixin (Amsellem et al 2014). Interestingly, meosin and stress-induced phosphoprotein-1, which were both downregulated in Δ43 hearts and not in A57G hearts, have been recently suggested to be possible sero-diagnostic markers of psoriasis (Maejima et al 2014).…”