Suboptimal B‐cell antigen receptor signaling activity in vivo elicits germinal center counterselection mechanisms

Abstract

Keywords: BCR r GC r Interclonal competition r Syk r Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionThe investigation of activation processes triggered by the B-cell antigen receptor has kept the scientific community busy for many years, in particular, understanding the spatial BCR organization Correspondence: Dr. Friedemann Kiefer e-mail: fkiefer@gwdg.de and the orchestration of kinase/phosphatase activity on resting versus activated B-cell membranes. Recent evidence argues that classical multimerization models for activation, postulating n ࣙ 2 BCRs to be cross-linked by antigen, have certain caveats and that receptor preoligomerization as well as protein islands might not only be a feature of T-cell surfaces, but also apply to B cells [1][2][3]. Besides this incomplete picture of BCR organization and autoinhibition, the importance of Syk as the central kinase conferring signaling potential to the B-cell antigen receptor is C 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.eji-journal.eu 604Sebastian Königsberger et al. Eur. J. Immunol. 2015. 45: 603-611 well established. Being inducibly recruited to phosphorylated Igα/β ITAMs that serve as allosteric kinase activatory sites [4], Syk phosphorylates a still growing list of signaling substrates, including SPL-65, PLCγ2, Btk, and Vav1. Thereby, Syk is central for the direct translation of receptor-mediated antigenic recognition into cellular responses such as Ca 2+ mobilization from the ER, actin remodeling, and initiation of de novo gene transcription [4][5][6]. The quality of the BCR signal is essential at every selection step during B-cell development, no matter if the signal is acute or tonic in nature [7]. Of note, the integrated signaling output during B-cell activation is a complex function of costimulatory activity and antigen receptor affinity/avidity, of which the latter is the result of RAG-mediated central recombination as well as AID-driven hypermutation in germinal centers (GCs) [8][9][10].We recently described a mouse model of constitutive Syk family kinase switch, in which we inserted a Zap-70 cDNA after the initiating ATG codon (exon 2) of the Syk gene, thereby disrupting endogenous Syk expression and driving Zap-70 from the endogenous Syk promoter (Syk ) [11]. In brief, the analysis of the B-cell compartment showed that developing Syk Zap-70/Zap-70 B cells less efficiently traverse the pre-BCR checkpoint, exhibit a bias toward marginal zone B-cell fate, and decrease BCR signaling potential due to inferior kinase activity. Of note, constitutive loss of Syk was shown to lead to a complete arrest of B-cell development at the immature stage [12,13]. Therefore, we were curious about what impact inducibly lowering Syk kinase levels, modeled here by monoallelic loxP site-flanked Syk in the presence of a knock-in Zap-70 allele that served as a hypoactive rescue allele in case of Syk deletion, would have on the propensity of B cells to participate in GC formation. To t...

Substrate stiffness regulates B‐cell activation, proliferation, class switch, and T‐cell‐independent antibody responses in vivo

“…The above points provide a mechanistic explanation for the enhanced activation of B cells on softer but not stiffer PDMS substrates, which we believe might also explain the similar observation in T cells. Moreover, the very recent BCR signaling pathway studies by Reth and his colleagues showed that cre-mediated Syk deletion in Sykflox/Zap-70 mouse primary B cells lowered the phosphorylation of Erk signaling molecule, but not the phosphorylation of Akt-mediated signaling, suggesting that Syk activation and Akt are not necessarily coupled signaling modulus in B-cell activation [41]. The contemporary data in this report echo back the early pioneer studies by Mond and Dintzis and their colleagues showing that anti-Ig (anti-IgM antibody as surrogate antigen) conjugated to dextran will be 1000-fold more potent to drive in vitro B-cell expansion than unconjugated soluble anti-Ig [42], suggesting that stiffness feature of the substrates presenting the antigens would dramatically affect the mutagenicity of these antigens to drive B-cell expansion.…”

“…The above points provide a mechanistic explanation for the enhanced activation of B cells on softer but not stiffer PDMS substrates, which we believe might also explain the similar observation in T cells. Moreover, the very recent BCR signaling pathway studies by Reth and his colleagues showed that cre‐mediated Syk deletion in Sykflox/Zap‐70 mouse primary B cells lowered the phosphorylation of Erk signaling molecule, but not the phosphorylation of Akt‐mediated signaling, suggesting that Syk activation and Akt are not necessarily coupled signaling modulus in B‐cell activation .…”

Checkpoints that control B cell development

“…By contrast, bone marrow-derived pHSCs give rise to shorter-lived BIb cells, which have homing properties that are similar to BIa cells. Most importantly, bone marrow develops BII-type B lymphocytes, which organize the B cell follicles in spleen and lymph nodes (85,86 Any high-affinity, autoreactive B cells that emerge as accidental products of the hypermutation process during this affinity maturation of better-fitting antibodies should be silenced and eliminated to avoid autoimmune disease (89)(90)(91)(92). Again, the microenvironments mediating this postulated negative selection of autoantibodies need further investigation.…”