Flow-induced reorganization of laminin-integrin networks within the endothelial basement membrane uncovered by proteomics

ABSTRACT: Objective The vessel wall is continuously exposed to hemodynamic forces generated by blood flow. Endothelial mechanosensors perceive and translate mechanical signals via cellular signaling pathways into biological processes that control endothelial development, phenotype and function. Here, we aim to unravel the molecular mechanisms underlying endothelial mechanosensing. Approach and results We applied a quantitative mass spectrometry approach combined with cell surface chemical footprinting to assess the hemodynamic effects on the endothelium on a system-wide level. These studies revealed that of the >5000 quantified proteins 104 were altered, which were highly enriched for extracellular matrix proteins and proteins involved in cell-matrix adhesion. Cell surface proteomics furthermore indicated that LAMA4 was proteolytically processed upon flow-exposure, which corresponded to the decreased LAMA4 mass observed on immunoblot. Immunofluorescence microscopy studies highlighted that the endothelial basement membrane was drastically remodeled upon flow exposure. We observed a network-like pattern of LAMA4 and LAMA5, which corresponded to the localization of laminin-adhesion molecules ITGA6 and ITGB4. Furthermore, the adaptation to flow-exposure did not affect the inflammatory response to tumor necrosis factor α, indicating that inflammation and flow trigger fundamentally distinct endothelial signaling pathways with limited reciprocity and synergy. Conclusions Taken together, this study uncovers the blood flow-induced remodeling of the basement membrane and stresses the importance of the subendothelial basement membrane in vascular homeostasis.

INSTRUMENT(S): Orbitrap Fusion Lumos, Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Endothelial Cell

SUBMITTER: Eelke Béguin  

LAB HEAD: Maartje van den Biggelaar

PROVIDER: PXD014526 | Pride | 2020-04-29

REPOSITORIES: Pride