Proline-rich tyrosine kinase 2 (Pyk2) mediates vascular endothelial-cadherin-based cell-cell adhesion by regulating beta-catenin tyrosine phosphorylation.
Vascular endothelial-cadherin (VE-cadherin) controls endothelial cell-cell adhesion and preserves endothelial integrity. In order to maintain endothelial barrier function, VE-cadherin function is tightly regulated through mechanisms that involve protein phosphorylation and cytoskeletal dynamics. Here, we show that loss of VE-cadherin function results in intercellular gap formation and a drop in electrical resistance of monolayers of primary human endothelial cells. Detailed analysis revealed that loss of endothelial cell-cell adhesion, induced by VE-cadherin-blocking antibodies, is preceded by and dependent on a rapid activation of Rac1 and increased production of reactive oxygen species. Moreover, VE-cadherin-associated beta-catenin is tyrosine-phosphorylated upon loss of cell-cell contact. Finally, the redox-sensitive proline-rich tyrosine kinase 2 (Pyk2) is activated and recruited to cell-cell junctions following the loss of VE-cadherin homotypic adhesion. Conversely, the inhibition of Pyk2 activity in endothelial cells by the expression of CRNK (CADTK/CAKbeta-related non-kinase), an N-terminal deletion mutant that acts in a dominant negative fashion, not only abolishes the increase in beta-catenin tyrosine phosphorylation but also prevents the loss of endothelial cell-cell contact. These results implicate Pyk2 in the reduced cell-cell adhesion induced by the Rac-mediated production of ROS through the tyrosine phosphorylation of beta-catenin. This signaling is initiated upon loss of VE-cadherin function and is important for our insight in the modulation of endothelial integrity.
van Buul
Jaap D
JD
Sanquin Research and Landsteiner Laboratory, Academic Medical Center, University of Amsterdam, 1066 CX Amsterdam, The Netherlands.
Anthony
Eloise C
EC
Fernandez-Borja
Mar
M
Burridge
Keith
K
Hordijk
Peter L
PL
D020727:ADP-Ribosylation Factors / Q000737:chemistry / Q000502:physiology*; D000256:Adenoviridae / Q000235:genetics; D015703:Antigens, CD; D015820:Cadherins / Q000378:metabolism*; D002448:Cell Adhesion; D002450:Cell Communication; D002478:Cells, Cultured; D003598:Cytoskeletal Proteins / Q000737:chemistry / Q000378:metabolism*; D003599:Cytoskeleton / Q000378:metabolism; D017097:Electric Impedance; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D051418:Focal Adhesion Kinase 2; D020690:GTPase-Activating Proteins / Q000737:chemistry / Q000502:physiology*; D017353:Gene Deletion; D005799:Genes, Dominant; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D009154:Mutation; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry; D010539:Permeability; D010766:Phosphorylation; D011392:Proline / Q000737:chemistry; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D017382:Reactive Oxygen Species; D015398:Signal Transduction; D013997:Time Factors; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*; D051176:beta Catenin; D020830:rac1 GTP-Binding Protein / Q000378:metabolism; D000099152:Cadherin 5
D020727:ADP-Ribosylation Factors; D015703:Antigens, CD; D015820:Cadherins; D003598:Cytoskeletal Proteins; D051418:Focal Adhesion Kinase 2; D020690:GTPase-Activating Proteins; D010455:Peptides; D011392:Proline; D011505:Protein-Tyrosine Kinases; D017382:Reactive Oxygen Species; D015534:Trans-Activators; D014443:Tyrosine; D042461:Vascular Endothelial Growth Factor A; D051176:beta Catenin; D020830:rac1 GTP-Binding Protein; D000099152:Cadherin 5; C495270:CTNNB1 protein, human; C502403:FAT1 protein, human; C410332:ASAP2 protein, human