Cytoskeletal Regulation of Lung Endothelial Pathobiology

DESCRIPTION (provided by applicant): This PPG renewal (years 11-15) remains focused on the lung vascular endothelium as a dynamic cellular barrier which is perturbed in inflammatory lung injury. The thrust of our work has elucidated the essential role of the endothelial cell (EC) cytoskeleton in vascular barrier regulation (disruption and restoration) as well as in diverse lung vascular processes (angiogenesis, apoptosis, etc). In this renewal, our outstanding group of gifted PPG scientists will continue to focus on the EC cytoskeleton as an essential participant in lung vascular barrier regulation. We will utilize physiologically- and clinically-relevant stimuli, incorporate genetically engineered murine models of inflammatory lung injury and explore genomically-driven protein targets. Project #1 will utilize state-of-the-art structure/ function studies of the non-muscle Ser/Thr myosin light chain kinase (MLCK) isoforms to extend our understanding of this exciting multi-functional protein as a molecular target in lung barrier regulation. As EC barrier-regulatory function is determined by cortical actin remodeling, Project #2 will provide novel information regarding the role of key actin-binding proteins (nmMLCK, cortactin, alpha actinin) and their cortical cytoskeletal linkages to junctional protein complexes. Project #3 examines the effects of mechanical stress (cyclic stretch) on EC signaling to the cytoskeleton in the context of barrier dysfunction/restoration via complex VEGF- and GTPase-driven actin rearrangement and focal adhesion remodelling. Project #4 will continue to provide its detailed examination of the NADPH oxidase as a target for regulation by the EC cytoskeleton. Project #5 will focus on the cascade of signaling to the actomyosin cytoskeleton evoked when c-Met, a tyrosine kinase receptor, is ligated by hepatocyte growth factor, a major barrier-regulatory pathway with strong potential for receptor transactivation. Supported by four highly interactive Cores (Administration, Tissue Culture/BioMechanical, Animal Models of Disease, Biophysical Imaging), we will utilize state-of-the-art biochemical, biophysical and molecular/genomic approaches that will not only likely provide the deepest understanding of the EC cytoskeleton and EC barrier regulation to date, but translationally define the role of EC cytoskeleton in critical biologic processes relevant to inflammatory lung injury. We anticipate our work will quickly facilitate development of new strategies and targets to limit the adverse effects of the injured pulmonary circulation.