Endothelial integrins that bind to provisional extracellular matrix (ECM) have been extensively studied, while the role of integrins involved in adhesion to vascular basement-matrix(BM), such as laminins, is less clear. Laminin-binding integrins such as α6β1 and α3β1 are considered important exclusively for the process of endothelial tube stabilization and their role in regulating sprouting angiogenesis is uncertain. We demonstrate that α6 integrin is up-regulated by angiogenic factors in endothelial cells (EC) and is high expressed in the angiogenic vasculature. In EC, where α6 integrin is down-regulated, defects in the ability to migrate and form tubular structures on laminin-containing matrix are prominently displayed. However, when similar experiments are performed on ECM ligands other than laminin, the α6 integrin down-regulation did not modify the EC response. Moreover, our results provide evidence that α6 integrin plays an unexpected role in adult pathological angiogenesis, showing that blockade of α6 integrin inhibits vessels formation both in CAM and in mammalian tumor transgenic model. The reduced tumor vessel diameter, together with the relative abundance of pericytes in mice treated with anti-α6 integrin antibody, suggest that the increase of α6 on EC contribute to the morphological and functional defects described in tumoral vessels. Furthermore, we observe that α6 integrin localizes to basal surface of EC and specifically in podosomes. Podosomes are specialized plasma-membrane microdomains that combine adhesive and proteolytic activities to spatially restricted sites of matrix degradation. We demonstrate the presence of α6 but not α3 integrin in the ring structure that surrounds the podosome core, both in EC in vitro and in aortic vessels in vivo. Moreover, laminin reduces α6 integrin localization to podosomes in a concentration-dependent manner. In conclusion, our results demonstrate that α6 is expressed on angiogenic EC in both culture and tumors, and it plays an important role in vascular sprouting and tumoral angiogenesis, regulating migration and proteolytic activity of EC.