Angiopoietin-1, Unlike Angiopoietin-2, Is Incorporated into the Extracellular Matrix via Its Linker Peptide Region

Angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2) affect angiogenesis differently during embryogenesis and tumorigenesis. In an attempt to understand the molecular basis underlying the distinct roles of those two homologous molecules, we investigated the association of Ang-1 and Ang-2 with the extracellular matrix (ECM). TA3 murine mammary carcinoma (TA3) and Lewis lung carcinoma cells expressing v5 epitope-tagged Ang-1 and Ang-2 were used in our studies. The results indicated that Ang-1 is secreted and incorporated into the ECM of the tumor cells, whereas Ang-2 is not associated with the ECM. The mutagenesis study indicated the domain that is responsible for the ECM association of Ang-1 is the linker peptide region between the coiled-coil and the fibrinogen-like domains. A weak binding between the coiled-coil domain of Ang-1 and the ECM was observed. Immunocytochemistry study revealed a distinct ECM distribution pattern of Ang-1, which is quite different from that of fibronectin, laminin, and collagen types I and IV. The ECM-associated Ang-1 proteins are released, and Tie-2 receptors are phosphorylated upon the adhesion of human umbilical vein endothelial cells. Implications of the difference in the ECM association of Ang-1 and Ang-2, which are related to the regulation of angiopoietin activity and their roles in local versus distant angiogenesis during tumor metastasis, are discussed.