SUMMARYIntegrin cell adhesion receptors and fibronectin, one of their extracellular matrix ligands, have been demonstrated to be important for angiogenesis using functional perturbation studies and complete knockout mouse models. Here, we report on the roles of the 5 and v integrins, which are the major endothelial fibronectin receptors, in developmental angiogenesis. We generated an integrin 5-floxed mouse line and ablated 5 integrin in endothelial cells. Unexpectedly, endothelial-specific knockout of integrin 5 has no obvious effect on developmental angiogenesis. We provide evidence for genetic interaction between mutations in integrin 5 and v and for overlapping functions and compensation between these integrins and perhaps others. Nonetheless, in embryos lacking both 5 and v integrins in their endothelial cells, initial vasculogenesis and angiogenesis proceed normally, at least up to E11.5, including the formation of apparently normal embryonic vasculature and development of the branchial arches. However, in the absence of endothelial 5 and v integrins, but not of either alone, there are extensive defects in remodeling of the great vessels and heart resulting in death at ~E14.5. We also found that fibronectin assembly is somewhat affected in integrin 5 knockout endothelial cells and markedly reduced in integrin 5/v double-knockout endothelial cell lines. Therefore, neither 5 nor v integrins are required in endothelial cells for initial vasculogenesis and angiogenesis, although they are required for remodeling of the heart and great vessels. These integrins on other cells, and/or other integrins on endothelial cells, might contribute to fibronectin assembly and vascular development.

We report the application of quantitative mass spectrometry to identify plasma membrane proteins differentially expressed in melanoma cells with high vs. low metastatic abilities. Using stable isotope labeling with amino acids in culture (SILAC) coupled with nanospray tandem mass spectrometry, we identified CUB-domaincontaining protein 1 (CDCP1) as one such differentially expressed transmembrane protein. CDCP1 is not only a surface marker for cells with higher metastatic potential, but also functionally involved in enhancing tumor metastasis. Overexpression of CDCP1 also correlates with activation of Src. Pharmacological reagents, PP2 and Dasatinib, which block Src family kinase activation, blocked scattered growth of CDCP1-overexpressing cells in 3D Matrigel culture, suggesting that CDCP1 might function through the activation of Src-family kinases (SFKs). This hypothesis was further supported by mutational studies of CDCP1. Whereas wild-type CDCP1 enhances Src activation, point mutation Y734F abolishes in vitro dispersive growth in 3D culture and in vivo metastasis-enhancing activities of CDCP1. In addition, the Y734F mutation also eliminated enhanced Src activation. Thus, this work provides molecular mechanisms for the metastasis-enhancing functions of CDCP1. D espite significant improvement in treatments for cancer, most mortality is due to tumor metastasis to distant organs. Tumor metastasis is a complicated process, presumably involving tumor cell detachment and migration/invasion from the primary site (local invasion); intravasation, survival in the circulation, arrest, and extravasation from the circulation (systemic dissemination); and growth, survival, and angiogenesis at the distant organ sites (colonization) (1-4). It has become increasingly clear that tumor cells cooperate with environmental factors, including other cell types in the tumor, such as fibroblasts, macrophages, platelets, and endothelial cells, as well as extracellular matrix, to metastasize (5-7). On the other hand, many tumor-intrinsic factors have also been identified that promote (8-11) or inhibit (12-14) tumor metastasis, using expression microarrays and other large-scale profiling technologies such as copy-number variation and SNP arrays.Plasma membrane proteins mediate communications between tumor cells and their microenvironment, acting in a sense as the "antennae" through which cells sense their microenvironment and determine cellular outcomes, such as cell proliferation, migration, or apoptosis, in response to the combined stimuli present in the microenvironment. Some plasma membrane proteins have been well studied, such as receptors for growth factors, cytokines, and chemokines, and a number have been shown to play important roles during various stages of tumor progression and metastasis. In addition, several cell-extracellular matrix and cell-cell adhesion molecules have been shown in different systems to contribute to tumor metastasis. Examples include up-regulation of integrin αV and CD44 and reduction of E-cadherin. Howeve...

Fibronectin containing alternatively spliced EIIIA and EIIIB domains is largely absent from mature quiescent vessels in adults, but is highly expressed around blood vessels during developmental and pathological angiogenesis. The precise functions of fibronectin and its splice variants during developmental angiogenesis however remain unclear due to the presence of cardiac, somitic, mesodermal and neural defects in existing global fibronectin KO mouse models. Using a rare family of surviving EIIIA EIIIB double KO mice, as well as inducible endothelial-specific fibronectin-deficient mutant mice, we show that vascular development in the neonatal retina is regulated in an autocrine manner by endothelium-derived fibronectin, and requires both EIIIA and EIIIB domains and the RGD-binding α5 and αv integrins for its function. Exogenous sources of fibronectin do not fully substitute for the autocrine function of endothelial fibronectin, demonstrating that fibronectins from different sources contribute differentially to specific aspects of angiogenesis.Electronic supplementary materialThe online version of this article (doi:10.1007/s10456-017-9563-8) contains supplementary material, which is available to authorized users.