ABSTRACT:The critical role of lung endothelial cells (ECs) and the ARDS vascular endotype in ARDS pathobiology isbeing dramatically highlighted in the current global COVID-19 pandemic. This highly translational PPG renewalwill address the contribution of unchecked EC permeability to the devastating multi-organ failure and mortality ofARDS by providing a comprehensive understanding at the molecular and genomic level of vascular barrier reg-ulation and repair. This renewal maintains its focus on the critical role of the EC cytoskeleton in the pathobiologyof ARDS and ventilator-induced lung injury (VILI) and seeks to directly address the unmet need for FDA-approved ARDS pharmacotherapies that attenuate lung vascular permeability and inflammation. We propose4 highly clinically-relevant tightly-woven PPG Projects centered on specific lung EC target proteins/genes thatare involved in: i) the unchecked vascular permeability and injury in ARDS; ii) vascular responses to exces-sive mechanical stress in VILI; iii) contributing to the genetic basis for ARDS health disparities in Africandescent subjects; and iv) providing novel ARDS therapeutic opportunities. Thematic integration across allprojects includes functional characterization of ARDS-associated SNPs and CpG sites in PPG target genes andthe role of ROS in transcriptional and biochemical regulation of peripheral cytoskeletal remodeling formation ofcytoskeletal-driven lamellipodia and focal adhesion (FA) reorganization that promotes EC gap closure. Project#1 will utilize system biology approaches to define novel cytoskeletal regulation of EC barrier responses by themulti-functional non-muscle myosin light chain kinase isoform (nmMLCK) and its cytoskeletal-binding partnercortactin. Project #2 (a new PPG addition) extends our recent report that EC secretion of the damage-associatedmolecular pattern (DAMP) protein eNAMPT (nicotinamide phosphoribosyltransferase) is critical to ARDS in-flammatory permeability and injury via eNAMPT ligation of the Toll-like receptor 4 (TLR4). Project #3 will inter-rogate the novel regulation by integrin 4 (ITGB4) and kindlin2 of the bidirectional signaling between the cyto-skeleton and focal adhesion (FA) dynamics which promotes lamellipodial-mediated closure of inflammation-in-duced EC gaps. Project #4 (a new PPG Project) will elucidate interactions between key barrier-regulatory re-ceptors: sphingosine-1-phosphate (S1P) receptor 1 (S1PR1) S1PR3 and P-selectin which regulate leukocyterecruitment and lung vascular leak via ligation by P-selectin glycoprotein ligand 1 (PSGL1). Supported by fourhighly interactive cores woven into each PPG Project (Administrative Genome/Proteome Preclinical AnimalModel Biophysical Imaging) this PPG will continue to leverage the outstanding scientific environments at theUniversity of Arizona and University of Illinois and the outstanding PPG translational team of interactive basicand physician-scientist investigators. In addition to informing the genetic basis for ARDS health disparities ourwork directly addresses the urgent unmet needs for greater mechanistic understanding of lung EC barrier regu-lation and novel therapeutics that restore the integrity of the injured lung circulation to reduce ARDS mortality.