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SENCR is a human-specific, vascular cell-enriched long-noncoding RNA (lncRNA) that regulates vascular smooth muscle cell and endothelial cell (EC) phenotypes. The underlying mechanisms of action of SENCR in these and other cell types is unknown. Here, levels of SENCR RNA are shown to be elevated in several differentiated human EC lineages subjected to laminar shear stress. Increases in SENCR RNA are also observed in the laminar shear stress region of the adult aorta of humanized SENCR -expressing mice, but not in disturbed shear stress regions. SENCR loss-of-function studies disclose perturbations in EC membrane integrity resulting in increased EC permeability. Biotinylated RNA pull-down and mass spectrometry establish an abundant SENCR -binding protein, cytoskeletal-associated protein 4 (CKAP4); this ribonucleoprotein complex was further confirmed in an RNA immunoprecipitation experiment using an antibody to CKAP4. Structure–function studies demonstrate a noncanonical RNA-binding domain in CKAP4 that binds SENCR . Upon SENCR knockdown, increasing levels of CKAP4 protein are detected in the EC surface fraction. Furthermore, an interaction between CKAP4 and CDH5 is enhanced in SENCR -depleted EC. This heightened association appears to destabilize the CDH5/CTNND1 complex and augment CDH5 internalization, resulting in impaired adherens junctions. These findings support SENCR as a flow-responsive lncRNA that promotes EC adherens junction integrity through physical association with CKAP4, thereby stabilizing cell membrane-bound CDH5.

Endothelial dysfunction, characterized by a decrease of nitric oxide (NO) bioavailability in the vessel wall, plays a crucial role in the pathogenesis of atherosclerosis. Oxidative stress due to increased reactive oxygen species (ROS) is implicated in endothelial dysfunction associated with diabetes. However the molecular mechanisms by which oxidative stress causes endothelial dysfunction remain incompletely understood. Blood flow, which generates fluid shear stress acting on endothelium, is the most potent physiological stimulus for NO production through endothelial NO synthase (eNOS) activation. Here we report that hyperglycemia and oxidative stress impairs fluid shear stress signal transduction and eNOS activation in endothelium. We found that the exposure of endothelial cells (ECs) to ROS generators such as menadione and xanthine/xanthine oxidase inhibited laminar flow-mediated Akt and eNOS phosphorylation and activation in human endothelial cells. Moreover, ECs pre-exposed to high glucose that generates ROS production in ECs, failed to respond to laminar flow for Akt/eNOS phosphorylation and activation. Consequently NO production from ECs in response to laminar flow was attenuated by high glucose treatment. Mechanistically, we observed that hyperglycemia and oxidative stress altered endothelial adherens junction integrity, manifested by the alternation of the localization of cell-cell junction molecules vascular endothelial cadherin (VE-cadherin) and beta-catenin. Silencing VE-cadherin or beta-catenin with small interference RNA also inhibited laminar flow-mediated signaling for eNOS activation, which mimics the effects of oxidative stress, suggesting that cell-cell junction integrity is critical for fluid shear stress signal transduction and eNOS activation. Collectively, our results demonstrate that hyperglycemia and oxidative stress impair laminar flow-mediated eNOS activation through the alternation of endothelial junction integrity and fluid shear stress signal transduction. Our findings suggest a novel mechanism whereby oxidative stress induces diabetes-associated endothelial dysfunction.