Annexin A8 promotes VEGF-A driven endothelial cell sprouting

The physiological and pathological process of angiogenesis relies on orchestrated endothelial cell (EC) adhesion, migration and formation of new vessels. Here we report that human umbilical vein endothelial cells (HUVECs) deficient in Annexin A8 (AnxA8), a member of the annexin family of CA²⁺- and membrane binding proteins, are strongly deficient in their ability to sprout in response to vascular endothelial growth factor (VEGF)-A, and are strongly impaired in their ability to migrate and adhere to β1 integrin-binding extracellular matrix (ECM) proteins. We find that these cells are defective in the formation of complexes containing the tetraspanin CD63, the main VEGF-A receptor VEGFR2/KDR/Flk-1, and the β1 Integrin subunit, on the cell surface. We observe that upon VEGF-A activation of AnxA8-depleted HUVECs, VEGFR2/KDR/Flk-1 internalization is reduced, phosphorylation of VEGFR2/KDR/Flk-1 is increased, and the spatial distribution of Tyr577-phosphorylated focal adhesion kinase (pFAK577) is altered. We conclude that AnxA8 affects CD63/VEGFR2/KDR/Flk-1/β1 Integrin complex formation, leading to hyperactivation of the VEGF-A signal transduction pathway, and severely disturbed VEGF-A-driven angiogenic sprouting.