Developmental Regulation of B Lymphocyte Immune Tolerance Compartmentalizes Clonal Selection from Receptor Selection

AbstractB lymphocyte development is a highly ordered process that involves immunoglobulin gene rearrangements, antigen receptor expression, and a learning process that minimizes the development of cells with reactivity to self tissue. Two distinct mechanisms for immune tolerance have been defined that operate during early bone marrow stages of B cell development: apoptosis, which eliminates clones of cells, and receptor editing, which spares the cells but genetically reprograms their autoreactive antigen receptors through nested immunoglobulin L chain gene rearrangements. We show here that sensitivity to antigen-induced apoptosis arises relatively late in B cell development and is preceded by a functionally distinct developmental stage capable of receptor editing. This regulation compartmentalizes clonal selection from receptor selection

Functional immunoglobulin transgenes guide ordered B-cell differentiation in Rag-1-deficient mice

We have examined the regulatory role of the individual components of the immunoglobulin antigen receptor in B-cell development by transgenic complementation of Rag-1 deficient (Rag-1⁻) mice. Complementation with a membrane µ heavy chain (µHC) gene allows progression of developmentally arrested Rag-1⁻ pro-B-cells to the small pre-B cell stage, whereas the introduction of independently integrated µHC and κ light chain (κLC) transgenes promotes the appearance of peripheral lymphocytes which, however, remain unresponsive to external stimuli. Complete reconstitution of the B-cell lineage and the emergence of functionally nature Rag-1⁻ peripheral B cells is achieved by the introduction of cointegrated heavy and light chain transgenes encoding an anti-H-2^k antibody. This experimental system demonstrates the competence of the µHC and κLC to direct and regulate the sequential stages of B-cell differentiation, defines the time at which negative selection of self-reactive B cells occurs, and shows that elimination of these cells occurs equally well in the absence of Rag-1 as in its presence. These data also support the hypothesis that Rag-1 directly participates in the V(D)J recombination process

Rearrangement of Mouse Immunoglobulin Kappa Deleting Element Recombining Sequence Promotes Immune Tolerance and Lambda B Cell Production

SummaryThe recombining sequence (RS) of mouse and its human equivalent, the immunoglobulin (Ig) kappa deleting element (IGKDE), are sequences found at the 3′ end of the Ig kappa locus (Igk) that rearrange to inactivate Igk in developing B cells. RS recombination correlates with Ig lambda (Igλ) light (L) chain expression and likely plays a role in receptor editing by eliminating Igk genes encoding autoantibodies. A mouse strain was generated in which the recombination signal of RS was removed, blocking RS-mediated Igk inactivation. In RS mutant mice, receptor editing and self-tolerance were impaired, in some cases leading to autoantibody formation. Surprisingly, mutant mice also made fewer B cells expressing lambda chain, whereas λ versus κ isotype exclusion was only modestly affected. These results provide insight into the mechanism of L chain isotype exclusion and indicate that RS has a physiological role in promoting the formation of λ L chain-expressing B cells

Enforced Bcl-2 Expression Inhibits Antigen-mediated Clonal Elimination of Peripheral B Cells in an Antigen Dose–dependent Manner and Promotes Receptor Editing in Autoreactive, Immature B Cells

The mechanisms that establish immune tolerance in immature and mature B cells appear to be distinct. Membrane-bound autoantigen is thought to induce developmental arrest and receptor editing in immature B cells, whereas mature B cells have shortened lifespans when exposed to the same stimulus. In this study, we used Eμ–bcl-2-22 transgenic (Tg) mice to test the prediction that enforced expression of the Bcl-2 apoptotic inhibitor in B cells would rescue mature, but not immature, B cells from tolerance induction. To monitor tolerance to the natural membrane autoantigen H-2Kb, we bred 3–83μδ (anti-Kk,b) Ig Tg mice to H-2b mice or to mice expressing transgene-driven Kb in the periphery. In 3–83μδ/bcl-2 Tg mice, deletion of autoreactive B cells induced by peripheral Kb antigen expression in the liver (MT-Kb Tg) or epithelia (KerIV-Kb Tg), was partly or completely inhibited, respectively. Furthermore, Bcl-2 protected peritoneal B-2 B cells from deletion mediated by acute antigen exposure, but this protection could be overcome by higher antigen dose. In contrast to its ability to block peripheral self-tolerance, Bcl-2 overexpression failed to inhibit central tolerance induced by bone marrow antigen expression, but instead, enhanced the receptor editing process. These studies indicate that apoptosis plays distinct roles in central and peripheral B cell tolerance

An immunoglobulin Cκ-reactive single chain antibody fusion protein induces tolerance through receptor editing in a normal polyclonal immune system

Understanding immune tolerance mechanisms is a major goal of immunology research, but mechanistic studies have generally required the use of mouse models carrying untargeted or targeted antigen receptor transgenes, which distort lymphocyte development and therefore preclude analysis of a truly normal immune system. Here we demonstrate an advance in in vivo analysis of immune tolerance that overcomes these shortcomings. We show that custom superantigens generated by single chain antibody technology permit the study of tolerance in a normal, polyclonal immune system. In the present study we generated a membrane-tethered anti-Igκ–reactive single chain antibody chimeric gene and expressed it as a transgene in mice. B cell tolerance was directly characterized in the transgenic mice and in radiation bone marrow chimeras in which ligand-bearing mice served as recipients of nontransgenic cells. We find that the ubiquitously expressed, Igκ-reactive ligand induces efficient B cell tolerance primarily or exclusively by receptor editing. We also demonstrate the unique advantages of our model in the genetic and cellular analysis of immune tolerance