Mechanical shear stress alters NOS3 interacted chromosome conformation [4C-seq]

The optimal expression of endothelial nitric oxide synthase (eNOS), the hallmark of endothelial homeostasis, is vital to vascular function. Dynamically regulated by various stimuli, eNOS expression is modulated at transcriptional, post-transcriptional, and post-translational levels. However, epigenetic modulations of eNOS, particularly through long non-coding RNAs (lncRNAs) and chromatin remodeling, remain to be explored. Here we identified an enhancer-associated lncRNA that enhances eNOS expression” (LEENE). Combining RNA-sequencing and chromatin conformation capture methods, we demonstrated that LEENE is co-regulated with eNOS and that its enhancer resides in proximity to eNOS promoter in endothelial cells (ECs). Deletion of LEENE enhancer locus decreases the LEEN-eNOS proximal association and eNOS mRNA level. Inhibition of LEENE using locked nucleic acids (LNA) decreases eNOS expression and monocyte adhesion to ECs, whereas overexpression of LEENE increases eNOS expression and eNOS-derived NO in ECs. Mechanistically, LEENE facilitates the recruitment of RNA Pol II to the eNOS promoter to enhance eNOS nascent RNA transcription. Our findings unravel a new layer in eNOS regulation and provide novel insights into cardiovascular regulation involving endothelial function. 4C-seq for the Endothelial Nitric oxide synthase (NOS3) in endothelial cells under mechanical wall shear stress.