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IgM exists as both a monomer on the surface of B cells and a pentamer secreted by plasma cells. Both pre-immune "natural" and antigen-induced "immune" IgM antibodies are important for protective immunity and for immune regulation of autoimmune processes by recognizing pathogens and self-antigens. Effector proteins interacting with the Fc portion of IgM, such as complement and complement receptors, have thus far been proposed but fail to fully account for the IgM-mediated protection and regulation. A major reason for this deficit in our understanding of IgM function seems to be lack of data on a long elusive Fc receptor for IgM (FcμR). We have recently identified a bona fide FcμR in both humans and mice. In this article we briefly review what we have learned so far about FcμR.

The T cell co-receptor, CD8, binds to the alpha 3 domain of HLA class I (Salter, R.D., R.J. Benjamin, P.K. Wesley, S.E. Buxton, T.P.J. Garrett, C. Clayberger, A.M. Krensky, A.M. Norman, D.R. Littman, and P. Parham. 1990. Nature [Lond.]. 345:41; Connolly, J.M., T.A. Potter, E.M. Wormstall, and T.H. Hansen. 1988. J. Exp. Med. 168:325; and Potter, T.A., T.V. Rajan, R.F. Dick II, and J.A. Bluestone. 1989. Nature [Lond.]. 337:73). To identify regions of CD8 that are important for binding to HLA class I, we performed a mutational analysis of the CD8 molecule in the immunoglobulin (Ig)-like variable domain. Our mutational analysis was based on our finding that using a cell-cell adhesion assay murine CD8 (Lyt-2) did not bind to human class I. Since the interaction of human CD8 with HLA class I is species specific, we substituted nonconservative amino acids from mouse CD8 and analyzed the ability of the mutated CD8 molecules expressed in COS 7 cells to bind HLA class I-bearing B lymphoblastoid cells, UC. Mutants with the greatest effect on binding were located in a portion of the molecule homologous to the first and second hypervariable regions of an antibody combining site. In addition, a panel of 12 anti-CD8 monoclonal antibodies were used to stain the 10 CD8 mutants, and amino acids that affected antibody binding were localized on the crystal structure of the Bence-Jones homodimer, REI. Support for an Ig-like structure of CD8 can be found in the pattern of substitutions affecting antibody binding. This work supports the similar tertiary structure of the CD8 alpha-terminal domain and an Ig variable domain.

The lymphocyte differentiation marker CD8 acts as a coreceptor with the T cell receptor (TCR) during recognition of peptide presented by major histocompatibility complex (MHC) class I molecules. The functions of CD8 in the TCR complex are thought to be signaling through the association of CD8 with the protein tyrosine kinase p56lck and adhesion to MHC class I through the alpha 3 domain. While the ability of the CD8 alpha/alpha homodimer to bind to classical MHC class I molecules has been shown, it is unclear whether CD8 can also bind nonclassical molecules. Of particular interest is human histocompatibility leukocyte antigen (HLA)-G which is expressed on placental cytotrophoblast cells. These cells do not express HLA-A, -B and -C molecules. In this report, we demonstrate that CD8 can bind to HLA-G. It is possible, therefore, that a cell bearing CD8 may interact with HLA-G-expressing cells.

An IgM-binding protein of approximately 60 kD has been identified on activated B cells, but not on resting and activated T cells, monocytes, or granulocytes. Here, we characterize this IgM-binding protein as a receptor for the Fc portion (CH3 and/or CH4 domains) of IgM molecules (Fc microR). The Fc microR can be expressed as a cell surface activation antigen throughout the pre-B and B cell stages in differentiation. Receptor expression is not directly linked with IgM production, as both mu- pre-B cells and isotype-switched B cells may express the Fc microR. The receptor molecules produced by both pre-B and B cells are identical in size and are characterized as an acidic sialoglycoprotein with O-linked, but no N-linked, oligosaccharide. The Fc microR is anchored to the surface of B-lineage cells via a glycosyl phosphatidylinositol linkage. The Fc microR is thus the third member of a family of Fc receptors expressed on B-lineage cells, and its preferential expression on activated B cells suggests a potential role in the response to antigens.

EBV can induce human B cells to proliferate, differentiate, and undergo transformation into continuously growing lymphoblastoid cell lines. The EBV responsiveness appears to be confined to a very limited subpopulation of B cells, the nature of which is still unclear. In these studies, we sorted tonsillar B cells on the basis of their expression of the early surface activation Ag, Bac-1, and compared their proliferative responses to EBV. Bac-1+ cells responded to EBV with a relatively high level of DNA synthesis, whereas the Bac-1- cells did not. Both large and small Bac-1+ cells were responsive to EBV and the responsiveness was unrelated to the level of Bac-1 immunofluorescence intensity. Bac-1+ cells were relatively enriched for surface IgM and IgD expression. When the Bac-1- population was enriched for IgM+ cells, the proliferative response was still significantly lower than that of the Bac-1+ population. B cells acquire the ability to bind IgM relatively late after activation, and this feature did not distinguish the EBV-responsive B cells. The results suggest B cells become responsive to EBV after an early activation signal.

When viable murine lymphocytes are incubated sequentially with a saturating amount of affinity-purified, rabbit anti-actin and highly conjugated FITC-goat anti-rabbit Ig, about 52% of mesenteric lymph node (MLN) lymphocytes and 36% of thymocytes exhibit a faint, but sharply punctate surface fluorescence. Cell surface actin (CSA) can be distinguished from staining of cytoplasmic actin in permeable cells, which are identified by their uptake of ethidium bromide. Staining of actin in ethidium bromide-permeable cells is 10-fold more intense than staining of actin on ethidium bromide-impermeable cells and is seen as uniformly fluorescent rings or crescents at the periphery of the cell and as dimmer, diffuse fluorescence centrally. Binding of rabbit anti-actin and goat anti-rabbit Ig to the lymphocyte cell surface is not mediated by Fc receptors; F(ab')2 fragments of these antibodies detect the same number of positive cells as do the intact molecules, and affinity-purified anti-KLH does not bind significantly. The cell surface stain, measured by flow cytometry or fluorescence microscopy, can be absorbed by pretreatment of the anti-actin with immobilized actin but not with IgG-Sepharose. Double-label experiments show that about 70% of the non-B cells and 30% of the MLN B cells bear detectable CSA. Although we have not ascertained the origin of CSA, we find that the number and brightness of cells exhibiting CSA cannot be increased by preincubating the cells with exogenous native skeletal muscle actin or with supernatant from dissociated MLN, indicating that there are no free binding sites for exogenous actin. The findings imply that either there is a developmentally expressed binding site(s) for actin, or that at various stages of development lymphocytes express a protein antigenically related to actin on their surface.

We have identified an IgM binding protein, a single chain polypeptide of Mr 60,000, that is expressed on the surface of B lymphocytes within 18 hr following their activation with phorbol myristate acetate. The IgM binding protein was also detected on fresh leukemic B cells from individuals with chronic lymphocytic leukemia, and the level of its expression was increased after phorbol myristate acetate activation. Resting and phorbol myristate acetate-activated T cells, monocytes, and polymorphonuclear leucocytes did not express detectable amounts of the IgM binding protein. The 60-kDa protein on activated human B cells could bind secreted IgM molecules of both mouse and human origin, as well as endogenous membrane-bound IgM molecules following their cross-linkage with anti-mu antibodies. The binding of soluble IgM molecules to the surface of activated B cells was also demonstrated by indirect immunofluorescence analysis.

We have produced a monoclonal antibody, Bac-1, that appears to identify a novel antigen on activated human B cells. The Bac-1 antigen can be detected between 8 to 16 hr, as well as transferrin receptors (T9), after activation of small resting B cells with phorbol myristic acetate, anti-IgM antibody, Staphylococcus aureus Cowan I, or Epstein-Barr virus. The expression of the Bac-1 antigen precedes that of IL 2 receptors (Tac-1). Peak expression of the Bac-1 antigen was observed on day 3 after activation, and decreased thereafter. The Bac-1 antigen was present on a minor subpopulation of relatively large B cells isolated from blood samples, and on "preactivated" B cells of heterogeneous size isolated from spleens and tonsils. It was not detected on bone marrow pre-B cells, blood small B cells, or plasma cells, nor was it expressed by resting or activated T cells or nonlymphoid cells. Certain B cell neoplasms and B lymphoblastoid cell lines were Bac-1+, but neoplastic cells of non-B lineage were Bac-1-. With immunoperoxidase staining, Bac-1+ cells were detected predominantly in the germinal centers of tonsil sections. The Bac-1 antigen on activated B cells was destroyed by protease treatment and was enhanced by neuraminidase treatment, suggesting that the Bac-1 antibody detects a cell surface molecule via an antigenic determinant which is partially obscured by neighboring sialic acid residues. The reactivity pattern of Bac-1 differs from the patterns of cellular reactivity reported for other monoclonal antibodies with specificity for activated human B cells.