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T-cell activation plays an essential role in the generation of the pulmonary inflammation that is manifest in allergic asthma. Optimal T-cell activation requires not only presentation of antigen with the major histocompatibility complex, but also concurrent signaling through costimulatory molecules. The costimulatory molecule SLAM (Signaling Lymphocytic Activation Molecule, CD150) is a glycoprotein expressed on activated lymphocytes and antigen-presenting cells. Disruption of the SLAM gene demonstrated that SLAM-induced signal transduction pathways regulate cytokine production by T helper (Th)2 cells and macrophages. Here we tested the postulate that the costimulatory molecule SLAM may be critical for allergic inflammation in a murine model. SLAM-deficient mice did not manifest allergen-induced bronchoalveolar lavage eosinophilia, increased serum IgE, or heightened airway responses compared with wild-type mice. Allergen-induced Th2 cytokines and Th1 cytokines were decreased in SLAM-deficient mice. These data support the concept that SLAM plays a crucial role in allergic responses.

Signaling lymphocyte activation molecule (SLAM, CD150, or SLAMF1) is a self-ligand receptor on the surface of activated T- and B-lymphocytes, macrophages, and dendritic cells (DCs). Here we examine the effect of SLAM/SLAM interactions on CD40L-induced CD40 signaling pathways in human DCs. CD40L-expressing L929 cells induced DCs to produce interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and IL-12, which was strongly inhibited by coexpression of SLAM on the surface of the L929 cells. Similarly, transfection of DCs with SLAM strongly reduced CD40L-induced IL-12 production. Furthermore, the negative effect of SLAM/SLAM interactions on CD40L-induced DC activation was also detected in the presence of lipopolysaccharide (LPS). LPS-induced IL-12 secretion, however, was not inhibited by SLAM engagement. CD40L-activated DCs affected by exposure to SLAM/SLAM engagement were impaired in their ability to induce differentiation of naive T lymphocytes into interferon-gamma (IFN-gamma)-producing T-helper 1 (Th1) effector cells. These inhibitory effects were not the result of a general unresponsiveness of DCs to CD40L, as SLAM/SLAM interactions did not prevent CD40L-induced up-regulation of CD83, CD86, or human leukocyte antigen (HLA)-DQ on the surface of DCs. Taken together, the results indicate that SLAM/SLAM interactions inhibit CD40-induced signal transduction in monocyte-derived dendritic cells, an effect that was not detectable in earlier studies using anti-SLAM monoclonal antibodies.

Human EAT-2 (SH2D1B) and SLAM-associated protein (SAP) (SH2D1A) are single SH2-domain adapters, which bind to specific tyrosine residues in the cytoplasmic tail of six signaling lymphocytic activation molecule (SLAM) (SLAMF1)-related receptors. Here we report that, unlike in humans, the mouse and rat Eat2 genes are duplicated with an identical genomic organization. The coding regions of the mouse Eat2a and Eat2b genes share 91% identity at the nucleotide level and 84% at the protein level; similarly, segments of introns are highly conserved. Whereas expression of mouse Eat2a mRNA was detected in multiple tissues, Eat2b was only detectable in mouse natural killer cells, CD8+ T cells, and ovaries, suggesting a very restricted tissue expression of the latter. Both the EAT-2A and EAT-2B coimmunoprecipitated with mouse SLAM in transfected cells and augmented tyrosine phosphorylation of the cytoplasmic tail of SLAM. Both EAT-2A and EAT-2B bind to the Src-like kinases Fyn, Hck, Lyn, Lck, and Fgr, as determined by a yeast two-hybrid assay. However, unlike SAP, the EAT-2 proteins bind to their kinase domains and not to the SH3 domain of these kinases. Taken together, the data suggest that both EAT-2A and EAT-2B are adapters that recruit Src kinases to SLAM family receptors using a mechanism that is distinct from that of SAP.

Platelet aggregation is a dynamic entity, capable of directing its own growth and stability via the activation of signaling cascades that lead to the expression and secretion of various secondary agonists. Here we show that the signaling pathways triggered during platelet aggregation include an intrinsic pro-thrombotic activity mediated by 2 homophilic adhesion molecules, CD84 and CD150 (SLAM [signaling lymphocyte activation molecule]), which are tyrosine phosphorylated in a platelet aggregation-dependent fashion. The 2 CD84/SLAM adapter proteins, SAP (SLAM-associated protein) and EAT-2 (EWS-activated transcript-2), were found in platelets; only SAP, however, was found to immunoprecipitate with tyrosine-phosphorylated SLAM. The immobilized extracellular domain of CD84 promoted microaggregate formation, while SAP-deficient platelets demonstrated defective spreading on immobilized CD84, demonstrating a functional role in platelets for SLAM family interactions. Finally, analysis of SLAM-deficient mice revealed an overall defect in platelet aggregation in vitro and a delayed arterial thrombotic process in vivo. The data indicate that signaling of the adhesion molecules in the SLAM family, activated by proximity during aggregation, further stabilize platelet-platelet interactions in thrombosis.

The primary manifestation of X-linked lymphoproliferative syndrome, caused by a dysfunctional adapter protein, signaling lymphocyte activation molecule-associated protein (SAP), is an excessive T cell response upon EBV infection. Using the SAP-/- mouse as a model system for the human disease, we compared the response of CD8+ T cells from wild-type (wt) and mutant mice to various stimuli. First, we observed that CD8+ T cells from SAP-/- mice proliferate more vigorously than those from wt mice upon CD3/CD28 cross-linking in vitro. Second, we analyzed the consequence of SAP deficiency on CTL effector function and homeostasis. For this purpose, SAP-/- and wt mice were infected with the murine gamma-herpesvirus 68 (MHV-68). At 2 wk postinfection, the level of viral-specific CTL was much higher in mutant than in wt mice, measured both ex vivo and in vivo. In addition, we established that throughout 45 days of MHV-68 infection the frequency of virus-specific CD8+ T cells producing IFN-gamma was significantly higher in SAP-/- mice. Consequently, the level of latent infection by MHV-68 was considerably lower in SAP-/- mice, which indicates that SAP-/- CTL control this infection more efficiently than wt CTL. Finally, we found that the Vbeta4-specific CD8+ T cell expansion triggered by MHV-68 infection is also enhanced and prolonged in SAP-/- mice. Taken together, our data indicate that SAP functions as a negative regulator of CD8+ T cell activation.

To improve our ability to identify hematopoietic stem cells (HSCs) and their localization in vivo, we compared the gene expression profiles of highly purified HSCs and non-self-renewing multipotent hematopoietic progenitors (MPPs). Cell surface receptors of the SLAM family, including CD150, CD244, and CD48, were differentially expressed among functionally distinct progenitors. HSCs were highly purified as CD150(+)CD244(-)CD48(-) cells while MPPs were CD244(+)CD150(-)CD48(-) and most restricted progenitors were CD48(+)CD244(+)CD150(-). The primitiveness of hematopoietic progenitors could thus be predicted based on the combination of SLAM family members they expressed. This is the first family of receptors whose combinatorial expression precisely distinguishes stem and progenitor cells. The ability to purify HSCs based on a simple combination of SLAM receptors allowed us to identify HSCs in tissue sections. Many HSCs were associated with sinusoidal endothelium in spleen and bone marrow, though some HSCs were associated with endosteum. HSCs thus occupy multiple niches, including sinusoidal endothelium in diverse tissues.

Ly108, a glycoprotein of the signaling lymphocytic activation molecule family of cell surface receptors expressed by T, B, NK, and APCs has been shown to have a role in NK cell cytotoxicity and T cell cytokine responses. In this study, we describe that CD4(+) T cells from mice with a targeted disruption of exons 2 and 3 of Ly108 (Ly108(DeltaE2+3)) produce significantly less IL-4 than wild-type CD4(+) cells, as judged by in vitro assays and by in vivo responses to cutaneous infection with Leishmania mexicana. Surprisingly, neutrophil functions are controlled by Ly108. Ly108(DeltaE2+3) mice are highly susceptible to infection with Salmonella typhimurium, bactericidal activity of Ly108(DeltaE2+3) neutrophils is defective, and their production of IL-6, IL-12, and TNF-alpha is increased. The aberrant bactericidal activity by Ly108(DeltaE2+3) neutrophils is a consequence of severely reduced production of reactive oxygen species following phagocytosis of bacteria. Thus, Ly108 serves as a regulator of both innate and adaptive immune responses.

Signaling lymphocyte activation molecule (SLAM), a glycoprotein expressed on activated lymphocytes and antigen-presenting cells, has been shown to be a coregulator of antigen-driven T cell responses and is one of the two receptors for measles virus. Here we show that T cell receptor-induced interleukin (IL)-4 secretion by SLAM(-/-) CD4(+) cells is down-regulated, whereas interferon gamma production by CD4(+) T cells is only slightly up-regulated. Although SLAM controls production of IL-12, tumor necrosis factor, and nitric oxide in response to lipopolysaccharide (LPS) by macrophages, SLAM does not regulate phagocytosis and responses to peptidoglycan or CpG. Thus, SLAM acts as a coreceptor that regulates signals transduced by the major LPS receptor Toll-like receptor 4 on the surface of mouse macrophages. A defective macrophage function resulted in an inability of SLAM(-/-) C57Bl/6 mice to remove the parasite Leishmania major. We conclude that the coreceptor SLAM plays a central role at the interface of acquired and innate immune responses.

The free Src homology 2 (SH2) domain protein SAP, encoded by the X-linked lymphoproliferative disease gene SH2D1A, controls signal transduction initiated by engagement of the SLAM-related receptors in T and NK cells. Here we demonstrate that SAP is required for phosphorylation of both SLAM and Ly9 in thymocytes and peripheral T cells. Furthermore, in vitro protein interaction studies and yeast two-hybrid analyses indicated that SAP binds directly to FynT and Lck. While SAP bound to both the SH3 domain and to the kinase domain of FynT, SAP bound solely to the kinase domain of Lck. The existence of a strong interaction between SAP and the SH3 domain of FynT prompted us to study the role of SAP in modulating the activity of FynT. In vitro addition of SAP to the autoinhibited form of FynT caused a large increase in FynT catalytic activity. By contrast, the SAP mutant R78E, which is unable to bind to the FynT SH3 domain, did not increase FynT activity and also displayed a reduced adaptor function upon transfection into T cells. Our results demonstrate that SAP is an adaptor that bridges SLAM and Ly9 with Src-like protein tyrosine kinases (PTKs), and has the ability to activate FynT.

SAP (signalling lymphocytic activation molecule (SLAM)-associated protein) is a T- and natural killer (NK)-cell-specific protein containing a single SH2 domain encoded by a gene that is defective or absent in patients with X-linked lymphoproliferative syndrome (XLP). The SH2 domain of SAP binds with high affinity to the cytoplasmic tail of the haematopoietic cell-surface glycoprotein SLAM and five related receptors. SAP regulates signal transduction of the SLAM-family receptors by recruiting SRC kinases. Similarly, the SAP-related proteins EAT2A and EAT2B are thought to control signal transduction that is initiated by SLAM-related receptors in professional antigen-presenting cells. In this review, we discuss recent findings on the structure and function of proteins of the SAP and SLAM families.

SAP (SLAM-associated protein) is a small lymphocyte-specific signalling molecule that is defective or absent in patients with X-linked lymphoproliferative syndrome (XLP). Consistent with its single src homology 2 (SH2) domain architecture and unusually high affinity for SLAM (also called CD150), SAP has been suggested to function by blocking binding of SHP-2 or other SH2-containing signalling proteins to SLAM receptors. Additionally, SAP has recently been shown to be required for recruitment and activation of the Src-family kinase FynT after SLAM ligation. This signalling 'adaptor' function has been difficult to conceptualize, because unlike typical SH2-adaptor proteins, SAP contains only a single SH2 domain and lacks other recognized protein interaction domains or motifs. Here, we show that the SAP SH2 domain binds to the SH3 domain of FynT and directly couples FynT to SLAM. The crystal structure of a ternary SLAM-SAP-Fyn-SH3 complex reveals that SAP binds the FynT SH3 domain through a surface-surface interaction that does not involve canonical SH3 or SH2 binding interactions. The observed mode of binding to the Fyn-SH3 domain is expected to preclude the auto-inhibited conformation of Fyn, thereby promoting activation of the kinase after recruitment. These findings broaden our understanding of the functional repertoire of SH3 and SH2 domains.

CD150 (signaling lymphocyte activation molecule [SLAM]) is a self-ligand cell surface glycoprotein expressed on T cells, B cells, macrophages, and dendritic cells. To further explore the role of CD150 signaling in costimulation and T(H)1 priming we have generated a panel of rat antimouse CD150 monoclonal antibodies. CD150 cell surface expression is up-regulated with rapid kinetics in activated T cells and lipopolysaccharide/interferon gamma (IFN-gamma)-activated macrophages. Anti-CD150 triggering induces strong costimulation of T cells triggered through CD3. DNA synthesis of murine T cells induced by anti-CD150 is not dependent on SLAM-associated protein (SAP, SH2D1A), because anti-CD150 induces similar levels of DNA synthesis in SAP(-/-) T cells. Antibodies to CD150 also enhance IFN-gamma production both in wild-type and SAP(-/-) T cells during primary stimulation. The level of IFN-gamma production is higher in SAP(-/-) T cells than in wild-type T cells. Anti-CD150 antibodies also synergize with interleukin 12 (IL-12) treatment in up-regulation of IL-12 receptor beta(2) mRNA during T(H)1 priming, and inhibit primary T(H)2 polarization in an IFN-gamma-dependent fashion. Cross-linking CD150 on CD4 T cells induces rapid serine phosphorylation of Akt/PKB. We speculate that this is an important pathway contributing to CD150-mediated T-cell proliferation.

Human CS1, also known as novel Ly9, 19A24, or CRACC, is a member of the immunoglobulin gene superfamily (IgSF) expressed on natural killer cells and other leukocytes. Here we describe the cloning of the mouse homologue of this gene. The mouse novel Ly9 gene is shown to encode a transmembrane protein composed of two extracellular immunoglobulin-like domains, a transmembrane region and an 88-amino acid cytoplasmic domain. Mouse novel Ly9 is structurally similar to the extracellular domains of CD84 and CD229 (Ly9). Both mouse and human novel Ly9 genes mapped close to the CD229gene in a region where other members of the CD150 family have also been mapped, and analysis of their genomic sequences showed that they have an identical intron/exon organization. Northern blot analysis revealed that the expression of mouse and human novel Ly9 was predominantly restricted to hematopoietic tissues, with the exception of testis. Here we show that SAP (SH2D1A), an adapter protein responsible for the X-linked lymphoproliferative disease, binds to the phosphorylated cytoplasmic tail of human but not mouse novel Ly9. Taken together, these data indicate that mouse novel Ly9 is a new member of the expanding CD150 family of cell surface receptors.

CD150 signaling lymphocytic activation molecule (SLAM), a T/B/dendritic cell surface glycoprotein, is a costimulatory receptor involved in T-cell activation and is also a receptor for measles virus. CD150-induced signal transduction is controlled by SAP/SH2D1A, the gene that is aberrant in X-linked lymphoproliferative disease and familial hemophagocytic lymphohistiocytosis. This report shows that CD150 colocalizes with the T-cell receptor (TCR) following CD3 triggering in human peripheral blood T cells and is rapidly and reversibly tyrosine phosphorylated on TCR cross-linking. The Src-like kinases Lck and Fyn phosphorylate tyrosine residues in the cytoplasmic tail of CD150. The results demonstrate that the SAP protein has 2 modes of binding to CD150. Binding to the motif Thr-Ile-Tyr281Ala-Gln-Val occurs in a phosphotyrosine-independent fashion and to the motif Thr-Val-Tyr327Ala-Ser-Val in a phosphotyrosine-dependent manner. Within both SAP binding motifs the threonine residue at position -2 to tyrosine is essential to stabilize the interaction irrespective of tyrosine phosphorylation, a feature unique to the SAP SH2 domain. A leucine residue, Leu278, further stabilizes nonphospho binding of SAP to Tyr281 of CD150. SAP blocking of the tyrosine phosphatase SHP-2 occurs primarily on Tyr281 of CD150 because SHP-2 requires both Tyr281 and Tyr327 for binding to CD150, and SAP binds to nonphosphorylated Tyr281. CD150 exhibits lateral mobility, segregating into intercellular contacts. The lateral mobility and homophilic clustering of CD150 between neighboring cells is not dependent on SAP/CD150 interaction.

The SLAM family of human genes currently consists of seven related members of the immunoglobulin superfamily, membrane-associated proteins, including CD150 (SLAM), CD244 (2B4), CD84, CD229 ( Ly-9), BLAME, CD48, and 19A. These genes are expressed to varying degrees in subsets of immune cells (T, B, natural killer, and myeloid cells) and may function as ligands or receptors. This set of genes, related to CD2 and CD58 on Chromosome (Chr) 1p98, are found clustered close together in the human genome on Chr 1q22. Four of these family members (CD150, CD244, CD84, CD229) contain conserved tyrosine motifs in their cytoplasmic tails that enable them to bind intracellular signaling molecules SAP and EAT-2. SAP is mutated in human X-linked lymphoproliferative disease (XLP), and studies in XLP patients have shown that improper signaling via molecules that bind SAP contributes to the disease. We have identified two new members of the SLAM family (SF), which we term SF2000 and SF2001, which are expressed in immune cells and map in the SLAM gene cluster. SF2001 does not contain SAP-binding motifs in its short cytoplasmic tail. SF2000, which is co-expressed with SAP in T cells, binds both SAP and EAT-2. The data suggest that signaling through SF2000, together with CD150, CD244, CD84, and CD229, is controlled by SAP and therefore contributes to the pathogenesis of XLP.

The T and natural killer (NK) cell-specific gene SAP (SH2D1A) encodes a 'free SH2 domain' that binds a specific tyrosine motif in the cytoplasmic tail of SLAM (CD150) and related cell surface proteins. Mutations in SH2D1A cause the X-linked lymphoproliferative disease, a primary immunodeficiency. Here we report that a second gene encoding a free SH2 domain, EAT-2, is expressed in macrophages and B lympho cytes. The EAT-2 structure in complex with a phosphotyrosine peptide containing a sequence motif with Tyr281 of the cytoplasmic tail of CD150 is very similar to the structure of SH2D1A complexed with the same peptide. This explains the high affinity of EAT-2 for the pTyr motif in the cytoplasmic tail of CD150 but, unlike SH2D1A, EAT-2 does not bind to non-phosphorylated CD150. EAT-2 binds to the phosphorylated receptors CD84, CD150, CD229 and CD244, and acts as a natural inhibitor, which interferes with the recruitment of the tyrosine phosphatase SHP-2. We conclude that EAT-2 plays a role in controlling signal transduction through at least four receptors expressed on the surface of professional antigen-presenting cells.

Human CD150 (SLAM) is a glycoprotein expressed on the surface of T, B, natural killer, and dendritic cells. The extracellular domain of CD150 is the receptor for measles virus and CD150 acts as a co-activator on T and B cells. We characterized the mouse and human CD150 genes, each of which comprises seven exons spanning approximately 32 kb. Mouse CD150 mRNA was detected in T cells and in most thymocyte subsets, except CD4-8- cells. Surprisingly, the CD4-8- thymocytes of CD3gammadeltanull mice, but not of Ragnull or severe combined immunodeficiency mice, expressed CD150. Whereas high levels of CD150 were found in Th1 cells, only small amounts were detectable in Th2 cells. CD150 expression was up-regulated upon in vitro activation of mouse T cells by anti-CD3. The complete mouse CD150 gene is highly homologous to its human orthologue in terms of nucleotide sequences and intron/exon organization. The human genomic sequences indicate that all isoforms detected so far have arisen from alternative splicing events. As judged by fluorescence in situ hybridization, mouse CD150 mapped to Chromosome (Chr) 1, band 1H2.2-2.3, and human CD150 was found on Chr 1q22. Human and mouse CD150 share sequence homologies with six other genes, five of which - CD84, CD229 (Ly-9), CD244 (2B4), CD48, and 19A - are localized in a 250-kb segment in close proximity to the human gene. Their location and their sequence similarities strongly suggest that the CD150 family of cell surface receptors arose via successive duplications of a common ancestral gene.

SAP, the gene that is altered or absent in the X-linked lymphoproliferative syndrome (XLP), encodes a small protein that comprises a single SH2 domain and binds to the cell-surface protein SLAM which is present on activated or memory T and B cells. Because defective NK cell activity also has been reported in XLP patients, we studied the SAP gene in NK cells. SAP was induced upon viral infection of SCID mice and shown to be expressed in NK cells by in vitro culturing in the presence of IL-2. Moreover, SAP was expressed in the NK cell lines YT and RNK 16. Because SLAM, the cell-surface protein with which SAP interacts, and 2B4, a membrane protein having sequence homologies with SLAM, also were found to be expressed on the surfaces of activated NK and T cell populations, they may access SAP functions in these populations. Whereas we found that 2B4 also binds SAP, 2B4-SAP interactions occurred only upon tyrosine phosphorylation of 2B4. By contrast, SLAM-SAP interactions were independent of phosphorylation of Y281 and Y327 on SLAM. As CD48, the ligand for 2B4, is expressed on the surface of Epstein-Barr virus (EBV)-infected B cells, it is likely that SAP regulates signal transduction through this pair of cell-surface molecules. These data support the hypothesis that XLP is a result of both defective NK and T lymphocyte responses to EBV. The altered responses may be due to aberrant control of the signaling cascades which are initiated by the SLAM-SLAM and 2B4-CD48 interactions.

SAP, the product of the gene mutated in X-linked lymphoproliferative syndrome (XLP), consists of a single SH2 domain that has been shown to bind the cytoplasmic tail of the lymphocyte coreceptor SLAM. Here we describe structures that show that SAP binds phosphorylated and nonphosphorylated SLAM peptides in a similar mode, with the tyrosine or phosphotyrosine residue inserted into the phosphotyrosine-binding pocket. We find that specific interactions with residues N-terminal to the tyrosine, in addition to more characteristic C-terminal interactions, stabilize the complexes. A phosphopeptide library screen and analysis of mutations identified in XLP patients confirm that these extended interactions are required for SAP function. Further, we show that SAP and the similar protein EAT-2 recognize the sequence motif TIpYXX(V/I).

In addition to triggering the activation of B- or T-cell antigen receptors, the binding of a ligand to its receptor at the cell surface can sometimes determine the physiological outcome of interactions between antigen-presenting cells, T and B lymphocytes. The protein SLAM (also known as CDw150), which is present on the surface of B and T cells, forms such a receptor-ligand pair as it is a self-ligand. We now show that a T-cell-specific, SLAM-associated protein (SAP), which contains an SH2 domain and a short tall, acts as an inhibitor by blocking recruitment of the SH2-domain-containing signal-transduction molecule SHP-2 to a docking site in the SLAM cytoplasmic region. The gene encoding SAP maps to the same area of the X chromosome as the locus for X-linked lymphoproliferative disease (XLP) and we found mutations in the SAP gene in three XLP patients. Absence of the inhibitor SAP in XLP patients affects T/B-cell interactions induced by SLAM, leading to an inability to control B-cell proliferation caused by Epstein-Barr virus infections.

The monoclonal antibodies, anti-T1 and anti-T3, both react with all human peripheral thymus-derived lymphocytes and with 10% of thymocytes; each, however, recognizes different cell surface structures. It was determined that the target antigen of anti-T is a 69 000 molecular weight cell surface glycoprotein and that the T3 antigen is a 19 000 mol. wt. glycoprotein.

The human cell surface antigen T3 is involved in several T lymphocyte specific functions, as determined by the effect of monoclonal antibodies (OKT3, anti-Leu-4, UCHT1) directed at this molecular structure. The main target antigen of these monoclonal antibodies is a glycoprotein of 20 kDa. It is associated with four, less predominant, structurally distinct glycoproteins of 25-28 kDa, 37 kDa and 44 kDa. Of these molecules only the 20-kDa T3 antigen could be labeled with the hydrophobic reagent 5-iodonaphthyl-1-azide (INA). Here we present evidence that the main 20-kDa T3 antigen is comprised of, in fact, two structurally different molecules. One of these is a glycoprotein with a protein backbone of 14 kDa, the other is an unglycosylated protein of 20 kDa. This unglycosylated protein is labeled specifically with INA. Additional evidence for the existence of two different 20-kDa T3 antigens is provided by studies using the enzymes endo-beta-N-acetylglucosaminidase H and endo-beta-N-acetylglucosaminidase F and the drug tunicamycin. We hypothesize that the specific susceptibility to labeling with INA of the unglycosylated 20-kDa T3 form reflects a positioning in the lipid bilayer different from that of the glycosylated 20-kDa T3 form.

A heteroantiserum raised to leukemic cells of a patient with non-T cell acute lymphoblastic leukemia (ALL) has been extensively absorbed with cells from a leukemic T cell line and an autologous B lymphoblastoid cell line to produce a common ALL antiserum (CALLA). CALLA is specific for leukemic cells of most patients with non-T cell ALL and chronic myelogenous leukemia (CML) in lymphoid blast crisis. It has been extensively tested on a wide variety of normal cells and is unreactive with them. CALLA identifies a surface glycoprotein having a m.w. of approximately 100,000 on reactive cell populations. In contrast, partially absorbed anti-ALL sera detect a similar glycoprotein band on CALLA-negative B and T cell lines. The glycoprotein identified by CALLA has been isolated and used as an immunogen. This new antiserum (C129) detects a 100,000-dalton glycoprotein not only on CALLA-positive cell populations but also on most CALLA-negative normal and malignant hematopoietic cells and on B and T cell lines. We conclude that there exists a family of 100,000-dalton glycoproteins that are present on a variety of normal, transformed, and malignant cells and that possess shared as well as unique antigenic regions. The expression of at least one of these antigens, detected by CALLA, may be tumor specific.

Here we characterize the T-cell-activating protein (TAP), an Ly-6 gene product involved in T cell activation, as a glycoprotein with a molecular weight of 10-12 kd under nonreducing conditions and 15-18 kd under reducing ones. Two of the three bands that are precipitated from metabolically labeled cells are expressed on the cell surface and can be recovered from the supernatants of cells treated with a phosphatidylinositol-specific phospholipase C. Thus TAP appears to be attached to the cell membrane via this lipid. Precisely the same anchorage is observed for the activating Thy-1 molecule, and is therefore of particular interest as a potentially novel linkage involved in membrane signal transduction.

Mo1, a phagocyte surface glycoprotein heterodimer, is involved in a number of phagocyte adhesion functions such as binding and ingestion of serum-opsonized particles, zymosan-induced degranulation, and superoxide generation. Deficiency of this antigen in humans has been associated with increased susceptibility to recurrent bacterial infections. The beta subunit of Mo1 is shared by another surface glycoprotein named LFA-1, which is involved in lymphocyte proliferation, cytolytic T cell, and natural killing activities. Two unrelated patients with Mo1 deficiency were found to be deficient in LFA-1 as well as in the common beta subunit. Investigation of lymphocyte functions in these two patients revealed normal mixed leukocyte culture-generated cytolytic T cell and natural killing activities and significantly reduced proliferative response to phytohemagglutinin. LFA-1-deficient cells also proliferated in response to soluble antigen and different alloantigens. These responses were partially blocked by anti-LFA-1 antibody. Whereas LFA-1 was undetectable by immunofluorescence and immunoprecipitation on the patients' resting T cells, significantly reduced (approximately 5% of normal) but detectable amounts of the heterodimeric LFA-1 antigen were found on mitogen and alloantigen-activated T cells. On granulocytes, Mo1 surface expression was also dependent on the state of cellular activation. The amount of surface Mo1 present on resting normal granulocytes increased by 3-10-fold following exposure to stimuli that induced degranulation, suggesting the presence of a major intracellular pool for this antigen. Analysis of subcellular fractions from granulocytes showed that intracellular Mo1 is located primarily in the specific granule fraction. Activated granulocytes had little or no increase in their surface expression of LFA-1 antigen. Deficient granulocytes had significantly increased numbers of Mo1 antigen expressed on the surface following stimulation with calcium ionophore (1 microM). However, the amount expressed continued to be significantly reduced compared with normal cells. Quantitation of surface Mo1 on granulocytes exposed to calcium ionophore (1 microM) showed that both parents in one family but only the mother in the other family had significantly reduced levels of Mo1, suggesting heterogeneity in the inheritance of this disorder. Whereas LFA-1 deficiency on lymphocytes was associated with normal alloantigen-induced cytolytic T cell and natural killing activities in these two patients, functions which were in part dependent on small amounts of detectable LFA-1 antigen, the Mo1 deficiency state led to significant defects in phagocyte adhesion functions. Hence, the clinical symptoms associated with this combined deficiency state reflect a more profound phagocyte than lymphocyte disorder.