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Lymphocyte migration from the blood into most secondary lymphoid organs is initiated by a highly selective adhesive interaction with the endothelium of specialized blood vessels known as high endothelial venules (HEV). The propensity of lymphocytes to migrate to particular lymphoid organs is known as lymphocyte homing, and the receptors on lymphocytes that dictate interactions with HEV at particular anatomical sites are designated "homing receptors". Based upon antibody blockade experiments and cell-type distribution studies, a prominent candidate for the peripheral lymph node homing receptor in mouse is the =90-kD cell surface glycoprotein (gp90 MEL) recognized by the monoclonal antibody MEL-14. Previous work, including sequencing of a cDNA encoding for this molecule, supports the possibility that gp90 uEL is a calcium-dependent lectin-like receptor. Here, we show that immunoaffinity-purified gp90 MEL interacts in a sugar-inhibitable manner with sites on peripheral lymph node HEV and prevents attachment of lymphocytes. Lymphocyte attachment to HEV in Peyer's patches, a gut-associated lymphoid organ, is not affected by g p 9 0 MEL. The results demonstate that gp90 MEL, as a lectin-like receptor, directly bridges lymphocytes to the endothelium. VUPHOCVTES move extensively throughout the body, migrating from the blood into lymphoid organs and back to the blood again via lymphatics. The overall process, known as lymphocyte recirculation, is an essential component of immune surveillance as it brings potentially responsive lymphocytes into contact with antigens sequestered within secondary lymphoid organs (reviewed in Butcher, 1986; Duijvestijn and Hamann, 1989; Ford, 1975; Stoolman, 1989; Woodruff et al., 1987; Yednock and Rosen, 1989). Entry of blood-borne lymphocytes into all secondary lymphoid organs, except the spleen, is initiated by a highly specific cell-cell recognition event between lymphocytes and the endothelial cells of specialized postcapillary venules found within these organs (Gowans, 1959; Gowans and Knight, 1964). These venules are characterized by having a cuboidal or high endothelial lining and are referred to as high endothelial venules (HEV). ~ The adhesive interactions between lymphocytes and HEV have been studied in vivo with short term recirculation studies (reviewed in Butcher et al., 1980) and in vitro with the highly validated Stamper-Woodruff adherence assay (Stamper and Woodruff, 1976), in which viable lymphocytes, overlaid onto cryostat-cut sections of lymphoid organs, selectively adhere to sectioned profiles of HEV. Three distinct adhesive specificities have been shown to ex1. Abbreviations used in this paper: HEV, high endothelial venules; PN, peripheral lymph node; PP, Peyer's patch. ist, one governing lymphocyte attachment to HEV in peripheral lymph nodes (PN), another for gut-associated Peyer's patches (PP), and a recently identified specificity for lungassociated lymph nodes (Butcher et al., 1980; Chin et al., 1983; Chin et al., 1984; Geoffroy et al., 1988). 2 It has been postulated that the endothelial cells of HEV bear cell surface ligands that are distinctive for particular lymphoid organs or anatomically related collections of lymphoid organs (Butcher and Weissman, 1984). The ligands are the heterophilic adhesion partners for a set of complementary lymphocyte receptors. Depending on a lymphocyte's developmental stage, class, subclass, and history of antigenic stimulation, the particular array of receptors expressed by the cell determines its ability to bind to and hence initiate extravasation across HEV in different anatomical sites (for review, see Butcher, 1986). "Homing receptor" is the designation applied to this set of lymphocyte adhesion receptors, reflecting their primary role in controlling the distinctive migratory or "homing" propensities of lymphocyte and lymphoma populations. The activity of these specific adhesion molecules appears to be supplemented by accessory molecules such as LFA-1, which act to strengthen the lymphocyte-HEV interaction without contributing to target organ specificity (Hamann et al., 1988; Pals et al., 1988). The use of this system of highly refined 2. Additional lymphocyte-endothelial adhesive specificities may regulate lymphocyte extravasation at sites of chronic inflammation such as inflamed joints (Jalkanen et al., 1986) and skin (Sackstein et al., 1988). The Rockefeller University Press, 0021-9525/89/1 ! /246317 $2.00 The Journal of Cell Biology, Volume 109, November 1989 2463-2469 2463 homing specificities is believed to lie in its potential to regulate the exact mix of functional subpopulations which populate each lymphoid organ such that immune responses initiated within are appropriate and maximally efficient for the particular anatomical site (Butcher, 1986). A number of candidates have been identified for HEVligands in the mouse (Streeter et al., 1988a,b) and for lymphocyte homing receptors in the mouse, rat, and human systems (Chin et al., 1986; Gallatin et al., 1983; Holzmann et al., 1989; Jalkanen et al., 1986; Rasmussen et al., 1985; Woodruff et al., 1987). In the rat, evidence exists that homing receptors are shed into lymph as soluble factors. One factor, termed high endothelial binding factor (HEBFLN), prevents lymphocyte attachment to PN HEV while having no effect on attachment to PP HEV when it is prereacted with tissue sections (Rasmussen et al., 1985). The other identified factor, HEBFpp, has the opposite activity (Chin et al., 1986). In the mouse, the most extensively studied candidate for a homing receptor is the '~90-kD (i.e., gp90 mL) glycoprotein detected on the surface of lymphocytes by the MEL-14 mAb (Gallatin et al., 1983). Lymphocytes pretreated with this antibody are incapable of attaching to PN HEV but bind at control levels to PP HEV. The cell surface level of gp90 uEL on various populations of normal lymphocytes and lymphomas correlates with their ability to bind to PN HEV (Gallatin et al., 1983; Reichert et al., 1983; Yednock et al., 1987a). In addition, our previous work has shown that a calcium-dependent, lectin-like receptor involved in PN homing is closely associated, and very likely identical, to gp90 mL (Yednoek et al., 1987b). In support of identity, cDNAs encoding gp90 ~sL predict a transmembrane glycoprotein with a calcium-dependent lectin domain at its extracellular amino terminus (Lasky et al., 1989; Siegelman et al., 1989). An EGFlike domain and two exact repeats of a complement regulatory domain comprise the remainder of the exlracellular region (Lasky et al., 1989; Siegelman et al., 1989). The present study confirms that gp90 M~L is an essential element of the PN homing receptor by showing that the purified antigen can interact with sites on PN HEV and prevent lymphocyte attachment in an organ-specific manner. Additionally, we provide the first direct evidence that specific sugars and polysaccharides can block the biological activity of the isolated receptor. Materials and Methods