Dietmar Vestweber

VE-Cadherin: The Major Endothelial Adhesion Molecule Controlling Cellular Junctions and Blood Vessel Formation

Vascular endothelial (VE)-cadherin is a strictly endothelial specific adhesion molecule located at junctions between endothelial cells. In analogy of the role of E-cadherin as major determinant for epithelial cell contact integrity, VE-cadherin is of vital importance for the maintenance and control of endothelial cell contacts. Mechanisms that regulate VE-cadherin-mediated adhesion are important for the control of vascular permeability and leukocyte extravasation. In addition to its adhesive functions, VE-cadherin regulates various cellular processes such as cell proliferation and apoptosis and modulates vascular endothelial growth factor receptor functions. Consequently, VE-cadherin is essential during embryonic angiogenesis. This review will focus on recent new developments in understanding the role of VE-cadherin in controlling endothelial cell contacts and influencing endothelial cell behavior by various outside-in signaling processes.

Junctional adhesion molecules (JAMs): more molecules with dual functions?

Junctional adhesion molecules (JAMs) are members of an immunoglobulin subfamily expressed by leukocytes and platelets as well as by epithelial and endothelial cells, in which they localize to cell-cell contacts and are specifically enriched at tight junctions. The recent identification of extracellular ligands and intracellular binding proteins for JAMs suggests two functions for JAMs. JAMs associate through their extracellular domains with the leukocyte β2 integrins LFA-1 and Mac-1 as well as with the β1 integrin α4β1. All three integrins are involved in the regulation of leukocyte-endothelial cell interactions. Through their cytoplasmic domains, JAMs directly associate with various tight junction-associated proteins including ZO-1, AF-6, MUPP1 and the cell polarity protein PAR-3. PAR-3 is part of a ternary protein complex that contains PAR-3, atypical protein kinase C and PAR-6. This complex is highly conserved through evolution and is involved in the regulation of cell polarity in organisms from Caenorhabditis elegans and Drosophila to vertebrates. These findings point to dual functions for JAMs: they appear to regulate both leukocyte/platelet/endothelial cell interactions in the immune system and tight junction formation in epithelial and endothelial cells during the acquisition of cell polarity.

The cell polarity protein ASIP/PAR-3 directly associates with junctional adhesion molecule (JAM)

The establishment and maintenance of cellular polarity are critical for the development of multicellular organisms. PAR (partitioning‐defective) proteins were identified in Caenorhabditis elegans as determinants of asymmetric cell division and polarized cell growth. Recently, vertebrate orthologues of two of these proteins, ASIP/PAR‐3 and PAR‐6, were found to form a signalling complex with the small GTPases Cdc42/Rac1 and with atypical protein kinase C (PKC). Here we show that ASIP/PAR‐3 associates with the tight‐junction‐associated protein junctional adhesion molecule (JAM) in vitro and in vivo. No binding was observed with claudin‐1, ‐4 or ‐5. In fibroblasts and CHO cells overexpressing JAM, endogenous ASIP is recruited to JAM at sites of cell–cell contact. Over expression of truncated JAM lacking the extracellular part disrupts ASIP/PAR‐3 localization at intercellular junctions and delays ASIP/PAR‐3 recruitment to newly formed cell junctions. During junction formation, JAM appears early in primordial forms of junctions. Our data suggest that the ASIP/PAR‐3–aPKC complex is tethered to tight junctions via its association with JAM, indicating a potential role for JAM in the generation of cell polarity in epithelial cells.

Angiopoietins assemble distinct Tie2 signalling complexes in endothelial cell–cell and cell–matrix contacts

The receptor tyrosine kinase Tie2, and its activating ligand Angiopoietin-1 (Ang1), are required for vascular remodelling and vessel integrity, whereas Ang2 may counteract these functions. However, it is not known how Tie2 transduces these different signals. Here, we show that Ang1 induces unique Tie2 complexes in mobile and confluent endothelial cells. Matrix-bound Ang1 induced cell adhesion, motility and Tie2 activation in cell–matrix contacts that became translocated to the trailing edge in migrating endothelial cells. In contrast, in contacting cells Ang1 induced Tie2 translocation to cell–cell contacts and the formation of homotypic Tie2–Tie2 trans-associated complexes that included the vascular endothelial phosphotyrosine phosphatase, leading to inhibition of paracellular permeability. Distinct signalling proteins were preferentially activated by Tie2 in the cell–matrix and cell–cell contacts, where Ang2 inhibited Ang1-induced Tie2 activation. This novel type of cellular microenvironment-dependent receptor tyrosine kinase activation may explain some of the effects of angiopoietins in angiogenesis and vessel stabilization.

MOLECULAR MECHANISMS THAT CONTROL LEUKOCYTE EXTRAVASATION : THE SELECTINS AND THE CHEMOKINES

Abstract Attachment of leukocytes to the blood vessel wall initiates leukocyte extravasation. This enables leukocytes to migrate to and accumulate at sites of tissue injury or infection where they execute host-defense mechanisms. A series of vascular cell adhesion molecules on leukocytes and on endothelial cells mediate leukocyte attachment to the endothelium in a stepwise process. A large panel of about 40 known human chemokines is able to specifically activate certain leukocytes and attract them to migrate across the endothelial barrier and within tissue. The specific combination of molecular signals provided by the diversity of cytokines, adhesion molecules, and chemokines regulates the specificity and selectivity of the recruitment of certain subpopulations of leukocytes in vivo. This review will focus on selectins and chemokines which initiate the cell contact and regulate activation and chemoattraction of leukocytes.

The gene defective in leukocyte adhesion deficiency II encodes a putative GDP-fucose transporter

Leukocyte adhesion deficiency II (LAD II) is characterized by the lack of fucosylated glycoconjugates, including selectin ligands, causing immunodeficiency and severe mental and growth retardation. No deficiency in fucosyltransferase activities or in the activities of enzymes involved in GDP-fucose biosynthesis has been found. Instead, the transport of GDP-fucose into isolated Golgi vesicles of LAD II cells appeared to be reduced. To identify the gene mutated in LAD II, we cloned 12 cDNAs from Caenorhabditis elegans, encoding multi-spanning transmembrane proteins with homology to known nucleotide sugar transporters, and transfected them into fibroblasts from an LAD II patient. One of these clones re-established expression of fucosylated glycoconjugates with high efficiency and allowed us to identify a human homolog with 55% identity, which also directed re-expression of fucosylated glycoconjugates. Both proteins were localized to the Golgi. The corresponding endogenous protein in LAD II cells had an R147C amino acid change in the conserved fourth transmembrane region. Overexpression of this mutant protein in cells from a patient with LAD II did not rescue fucosylation, demonstrating that the point mutation affected the activity of the protein. Thus, we have identified the first putative GDP-fucose transporter, which has been highly conserved throughout evolution. A point mutation in its gene is responsible for the disease in this patient with LAD II.

Adhesion and signaling molecules controlling the transmigration of leukocytes through endothelium

Summary:  Migration of leukocytes into tissue is a key element of innate and adaptive immunity. While the capturing of leukocytes to the blood vessel wall is well understood, little is known about the mechanisms underlying the actual transmigration of leukocytes through the vessel wall (diapedesis). Even a basic question such as whether leukocytes migrate through openings between adjacent endothelial cells (junctional pathway) or through single endothelial cells (transcellular pathway) is still a matter of intensive debate. It is generally accepted that both pathways exist; however, whether they are of equal physiological significance is unclear. Several endothelial adhesion and signaling molecules have been identified, most of them at endothelial cell contacts, which participate in leukocyte diapedesis. A concept is evolving suggesting that transendothelial migration of leukocytes is a stepwise process. Blocking or eliminating some of the different adhesion and signaling proteins results in very different effects, such as trapping of leukocytes above endothelial cell contacts, in between endothelial cells, or between the endothelium and the underlying basement membrane. Other proteins are involved in the opening of endothelial cell contacts and yet others in their maintenance providing the barrier for extravasating leukocytes. The various molecular players and the functional steps involved in diapedesis are discussed.

VE-PTP and VE-cadherin ectodomains interact to facilitate regulation of phosphorylation and cell contacts

VE‐cadherin is the essential adhesion molecule in endothelial adherens junctions, and the regulation of protein tyrosine phosphorylation is thought to be important for the control of adherens junction integrity. We show here that VE‐PTP (vascular endothelial protein tyrosine phosphatase), an endothelial receptor‐type phosphatase, co‐precipitates with VE‐cadherin, but not with β‐catenin, from cell lysates of transfected COS‐7 cells and of endothelial cells. Co‐precipitation of VE‐cadherin and VE‐PTP required the most membrane‐proximal extracellular domains of each protein. Expression of VE‐PTP in triple‐transfected COS‐7 cells and in CHO cells reversed the tyrosine phosphorylation of VE‐cadherin elicited by vascular endothelial growth factor receptor 2 (VEGFR‐2). Expression of VE‐PTP under an inducible promotor in CHO cells transfected with VE‐cadherin and VEGFR‐2 increased the VE‐cadherin‐mediated barrier integrity of a cellular monolayer. Surprisingly, a catalytically inactive mutant form of VE‐PTP had the same effect on VE‐cadherin phosphorylation and cell layer permeability. Thus, VE‐PTP is a transmembrane binding partner of VE‐cadherin that associates through an extracellular domain and reduces the tyrosine phosphorylation of VE‐cadherin and cell layer permeability independently of its enzymatic activity.

GDF-15 is an inhibitor of leukocyte integrin activation required for survival after myocardial infarction in mice

Inflammatory cell recruitment after myocardial infarction needs to be tightly controlled to permit infarct healing while avoiding fatal complications such as cardiac rupture. Growth differentiation factor-15 (GDF-15), a transforming growth factor-β (TGF-β)–related cytokine, is induced in the infarcted heart of mice and humans. We show that coronary artery ligation in Gdf15-deficient mice led to enhanced recruitment of polymorphonuclear leukocytes (PMNs) into the infarcted myocardium and an increased incidence of cardiac rupture. Conversely, infusion of recombinant GDF-15 repressed PMN recruitment after myocardial infarction. In vitro, GDF-15 inhibited PMN adhesion, arrest under flow and transendothelial migration. Mechanistically, GDF-15 counteracted chemokine-triggered conformational activation and clustering of β2 integrins on PMNs by activating the small GTPase Cdc42 and inhibiting activation of the small GTPase Rap1. Intravital microscopy in vivo in Gdf15-deficient mice showed that Gdf-15 is required to prevent excessive chemokine-activated leukocyte arrest on the endothelium. Genetic ablation of β2 integrins in myeloid cells rescued the mortality of Gdf15-deficient mice after myocardial infarction. To our knowledge, GDF-15 is the first cytokine identified as an inhibitor of PMN recruitment by direct interference with chemokine signaling and integrin activation. Loss of this anti-inflammatory mechanism leads to fatal cardiac rupture after myocardial infarction.

Expression and distribution of cell adhesion molecule uvomorulin in mouse preimplantation embryos

We have examined the synthesis and distribution of the cell adhesion molecule uvomorulin in mouse preimplantation embryos. Uvomorulin can already be detected on the cell surface of unfertilized and fertilized eggs but is not synthesized in these cells. Uvomorulin synthesis starts in late two-cell embryos and seems not to be correlated with the onset of compaction. The first signs of compaction are accompanied by a redistribution of uvomorulin on the surface of blastomeres. During compaction uvomorulin is progressively removed from the apical membrane domains of peripheral blastomeres. In compact morulae uvomorulin is no longer present on the outer surface of the embryo but is localized predominantly in membrane domains involved in cell-cell contacts of adjacent outer blastomeres. On inner blastomeres of compact morulae uvomorulin remains evenly distributed. This uvomorulin distribution once established during compaction is maintained and also found in the blastocyst: on trophectodermal cells uvomorulin localization is very similar to that in adult intestinal epithelial cells while uvomorulin remains evenly distributed on the surface of inner cell mass cells. The possible role of the redistribution of uvomorulin for the generation of trophectoderm and inner cell mass in early mouse embryos is discussed.