Two novel mAbs, LIA1/1 and VJ1/16, specifically recognized a 25-kD protein that was selectively localized at cellCcell junctions of EC, both in the primary formation of cell monolayers and when EC reorganized in the process of wound healing. CD81/TAPA-1 inhibited the movement of individual ECs, as determined by quantitative time-lapse video microscopy studies. Furthermore, mAbs against the CD151/PETA-3 molecule diminished the bio-THZ1 pace of EC invasion into collagen gels. In addition, these mAbs were able to increase the adhesion of EC to extracellular bio-THZ1 matrix proteins. Collectively these results show that CD81/TAPA-1 and CD151/PETA-3 tetraspan molecules are components of the endothelial lateral junctions implicated in the rules of cell motility, either directly or by modulation of the function of the connected integrin heterodimers. Intercellular adhesion constructions provide, by means of transmembrane proteins selectively localized at the sites of cellCcell contact, the physical strength necessary to build up solid cells interconnecting the cytoskeleton from the different cells. Junctional constructions will also be responsible bio-THZ1 for the polarization of particular cell types, determining different practical subdomains along the plasma membrane, each comprising a defined subset of proteins. Tight junctions, made up from the transmembrane protein occludin (Furuse et al., 1993) coupled to cytoplasmic proteins ZO-1, ZO-2, 7H6, cingulin, and symplekin (Keon et al., 1996; for review observe Schneeberger et al., 1992; Anderson et al., 1993; Citi 1993), are directly involved in restricting the lateral diffusion of proteins along the aircraft of the plasma membrane. Adherens junctions, created by different cadherins (examined in Takeichi, 1990; Geiger and Ayalon, 1992; Dejana 1996) linked to the actin cytoskeleton by catenins (Tsukita et al., 1992; Kemler 1993; Cowin and Burke, 1996), initiate cellCcell contacts, nucleate the formation of additional junctional constructions (Gumbiner et al., 1988), and regulate the manifestation of the genes involved in the polarized phenotype bio-THZ1 (McNeill et al., 1990; Marrs et al., 1995). Focal adhesions, in which integrins are the transmembrane adhesion moiety, are primarily responsible for adhesion to the extracellular matrix (Jockusch et al., 1995), which may be adequate for the establishment of some of the characteristics of a polarized cell phenotype (Drubin and Nelson, 1996). Additional junctional complexes like space junctions, made up by connexin oligomers (for review observe Goodenough et al., 1996), do not play a structural part in intercellular adhesion but metabolically couple cells inside a determinate cells. Intercellular contacts are responsible for the main function of endothelial cells like a selective permeable barrier between the bloodstream and the rest of cells along the body. Endothelial cell-to-cell adhesion also takes on the aforementioned general part of cell growth rate control (Caveda et al., 1996) and cells integrity maintenance. Growth control in endothelium has a great relevance in tumorigenesis, since angiogenesis is one of the main requisites for tumor progression and metastasis (Hanahan and Folkman, 1996). On the other hand, intercellular connections must be modulated by many different stimuli in order to finely regulate the permeability of the endothelial cells (EC)1 monolayer to plasma macromolecules and, in certain cells and inflammatory conditions, to defined subpopulations of leukocytes present in the bloodstream. Vascular endothelial (VE)-cadherin, an endothelium-specific member of the superfamily of cadherins, seems to be one of the main regulators of permeability in EC monolayers. VE-cadherin is definitely reversibly linked to actin cytoskeleton by catenins and its association with these proteins is rapidly controlled through phosphorylation on catenin tyrosine residues (Lampugnani et al., 1992; Dejana 1996). Additional adhesion molecules, such as CD31/PECAM (platelet-endothelial cell adhesion molecule), also localize at intercellular contact sites where it may play a functional part much like VE-cadherin. CD31 mediates both homophilic as well as heterophilic (CD31-v3) molecular relationships, and is involved in the leukocyte transmigration across the EC monolayer (examined in Newman 1997). Certain integrins, such as 21 and 51, have also been implicated in the maintenance of the EC monolayer integrity (Lampugnani et al., 1991). The tetraspan superfamily of proteins (TM4) comprises a group of molecules with four membrane-spanning domains, which have been implicated in several cellular functions, as rules of cell growth and differentiation, cell adhesion, and intracellular signaling (examined in Wright and Tomlison, 1994; Hemler et al., 1996; Maecker et al., 1997). In this study, we show the specific intercellular localization of CD9, CD81/TAPA-1 and CD151/PETA-3 tetraspan molecules in EC as well as their specific connection with 31 integrin. These tetraspan molecules appear to possess an important part in EC motility, likely by altering cellCmatrix adhesion. Materials Rabbit Polyclonal to CYSLTR2 and Methods Cells and Cell Cultures Human being EC from umbilical vein (HUVEC) were.