Secondary antibody responses are marked by faster kinetics improved antibody BINA affinity and a switch from IgM to other immunoglobulin isotypes most notably IgG compared with primary responses. IgG antibody responses. Figure Rabbit Polyclonal to KLF11. 4 The ITT promotes IgG antibody production. The ITT improves the competitiveness of IgG-switched cells To compare the performance of wild-type and ITT-mutant B cells directly in the same animals we bred mIgG1-YF BL/6 mice expressing ITT-mutant mIgG1b with Balb/c mice expressing wild-type mIgG1a. Owing to allelic exclusion B cells of heterozygous F1 animals expressed either wild-type (allotype a) BINA or YF-mutant (allotype b) mIgG1. Animals that had both allotypes in wild-type form served as controls. Allotype-specific analysis of IgG1 production in animals that were immunized with ovalbumin (OVA) in aluminium hydroxide (alum) showed that wild-type mIgG1a-expressing B cells contributed up to six times more to the production of soluble IgG1 than their mIgG1b-expressing ITT-mutant counterparts both during primary and secondary responses (Fig. 5a c grey graph). In control animals both allotypes of IgG1 were produced in equal amounts (Fig. 5b) and hence were present in an almost 1:1 ratio during the entire course of the experiment (Fig. 5c black graph). In conclusion these results reveal that ITT-mediated signal amplification increases the competitiveness of mIgG-expressing cells which facilitates their contribution to antibody production. Figure 5 The ITT augments the competitiveness of mIgG-expressing B cells. ITT signals promote plasma cell formation To test whether reduced antibody production by ITT-mutant B cells was linked to impaired plasma cell generation we determined the frequency of OVA-specific IgG1 antibody forming cells in heterozygous F1 BINA animals at day 21 of the recall response by means of allotype-specific ELISPOT assays (Fig. 6a). In accordance with the diminished IgG1b serum titres (Fig. 5) we observed an approximately fourfold reduction in the number of YF-mutant IgG1b-producing antibody forming cells compared with IgG1a-producing cells both in the bone marrow (Fig. 6a b d) as well as in the spleens of heterozygous BINA F1 animals (Fig. 6c d). We conclude that ITT signalling supports the generation of plasma cells from the memory cell compartment. Figure 6 The ITT promotes plasma cell generation. ITTs promote T-cell-independent activation of memory B cells The reactivation requirements of memory B cells particularly the need for T-cell help are not fully understood. However reactivation of memory B cells by antigen especially viruses can occur independently of BINA T cells24 25 26 27 yet seems to require specific niches in secondary lymphoid tissue28. To test whether ITT signalling contributes to the improved reactivation of IgG-switched memory B cells in the absence of T-cell help we employed a cell transfer approach in which wild-type and YF-mutant mIgG1-expressing memory B cells were transferred into gene in the mouse impairs reactivation of IgG-switched memory B cells corroborating the importance of the ITT-Grb2 interaction for efficient antibody recall responses17 29 The most salient signalling effect of ITT-mediated Grb2-recruitment into the BCR signalosome is the enhanced activation of phospholipase C-γ2 (PLC-γ2) concomitant with a greatly prolonged influx of Ca2+ across the plasma membrane. In line with this homoeostasis of B-cell memory relies on the expression of PLC-γ2 since its cell-type-specific ablation in mIgG1-expressing B cells causes reduced formation and survival of IgG1-switched memory B cells30. Furthermore in B cells the phosphatase calcineurin which controls the activation of transcription factor NF-AT is specifically required BINA for terminal differentiation into plasma cells31. Considering that the activity of calcineurin is stimulated by Ca2+/calmodulin it appears possible that ITT-mediated prolongation of mIgG-BCR-induced Ca2+ mobilization augments the activity of calcineurin thereby supporting the differentiation of IgG-switched B cells into plasma cells. Plasma cell differentiation is generally considered to be governed by two antagonizing groups of transcriptional regulators that either maintain the mature B-cell phenotype such as Pax5 and Bcl-6 or induce the plasma cell differentiation programme like Irf4 and Blimp-1 (ref. 32). Expression of either set represses the other one and elimination of Bcl-6.

Secondary antibody responses are marked by faster kinetics improved antibody BINA
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