Vascular endothelial cadherin (VE-cadherin) mediates contact inhibition of cell growth in quiescent endothelial cell layers. Searching for proteins that could be involved in VE-cadherin signaling, we found the cytosolic C-terminal Src kinase (Csk), a negative regulator of Src family kinases. We show that Csk binds via its SH2 domain to the phosphorylated tyrosine 685 of VE-cadherin. VE-cadherin recruits Csk to cell contacts and both proteins can be co-precipitated from cell lysates of transfected cells and endothelial cells. Association of VE-cadherin and Csk in endothelial cells increased with increasing cell density. CHO cells expressing the tyrosine replacement mutant VE-cadherin-Y685F grow to higher cell densities than cells expressing wild-type VE-cadherin. Overexpression of Csk in these cells under an inducible promoter inhibits cell proliferation in the presence and absence of VE-cadherin, but not in the presence of VE-cadherin-Y685F. Reduction of Csk expression by RNA interference enhances endothelial cell proliferation. Our results suggest that the phosphorylated tyrosine residue 685 of VE-cadherin and probably the binding of Csk to this site are involved in inhibition of cell growth triggered by cell density.