Integrin linked kinase (ILK) is an extracellular matrix (ECM) binding molecule which links the b1 integrin cytosolic domain. We investigated the relevance of ILK for endothelial cell and vessel formation using ILK (-/-) embryonic stem (ES) cells. We found that ILK deficiency results in strongly reduced and defective formation of vessel-like structures. To decipher potentially underlying signalling defect(s) we performed [Ca2+]i imaging experiments on sorted endothelial cells (ECs) from WT and ILK (-/-) EBs. We found that VEGF application (20ng/ml) led in 67.1 % of WT ECs to a [Ca2+]i increase whereas in ILK (-/-) ECs almost all cells (99.1%, n=90) did not respond. In analogy, also the EGF (20ng/ml) response was found to be absent in the large majority of ILK (-/-) ECs (1.7 %; n=59, 72.4%, n=21 WT). This indicates defective VEGF/EGF signaling due to reduced tyrosine kinase receptor activation. Since these receptors and their signalling pathways are mostly organized in caveolae, we performed time lapse microscopy using ILK (-/-) ECs transfected with a caveolin-1-EGFP fusion protein; this molecule plays a critical role for organizing caveolae-dependent signalling clusters. We revealed striking defects in the positioning of caveolin-1 at the plasma membrane; moreover the defects in caveolin-1 dynamics (the average velocity was significantly higher in WT (0.29±0.02 mm/s, n=41) than in ILK (-/-) (0.09±0.01 mm/s, n=42)) were found to be accompanied by a disrupted interaction between microtubules and cortical actin filaments. Thus, ILK is required as a scaffold to allow actin/microtubule interaction and to place caveolin-1 at the plasma membrane. The correct spatial positioning of caveolae is a critical pre-requisite for key signalling pathways in endothelial cells.