A strong humoral response to infection requires the collaboration of several hematopoietic cell types that communicate via antigen presentation, surface coreceptors and their ligands, and secreted factors. The proinflammatory cytokine IL-6 has been shown to promote the differentiation of activated CD4+ T cells into T follicular helper cells (TFH cells) during an immune response. TFH cells collaborate with B cells in the formation of germinal centers (GCs) during T cell–dependent antibody responses, in part through secretion of critical cytokines such as IL-21. In this study, we demonstrate that loss of either IL-6 or IL-21 has marginal effects on the generation of TFH cells and on the formation of GCs during the response to acute viral infection. However, mice lacking both IL-6 and IL-21 were unable to generate a robust TFH cell–dependent immune response. We found that IL-6 production in follicular B cells in the draining lymph node was an important early event during the antiviral response and that B cell–derived IL-6 was necessary and sufficient to induce IL-21 from CD4+ T cells in vitro and to support TFH cell development in vivo. Finally, the transcriptional activator Oct2 and its cofactor OBF-1 were identified as regulators of Il6 expression in B cells.

The transcriptional network regulating antibody-secreting cell (ASC) differentiation has been extensively studied, but our current understanding is limited. The mechanisms of action of known “master” regulators are still unclear, while the participation of new factors is being revealed. Here, we identify Zbtb20, a Bcl6 homologue, as a novel regulator of late B cell development. Within the B cell lineage, Zbtb20 is specifically expressed in B1 and germinal center B cells and peaks in long-lived bone marrow (BM) ASCs. Unlike Bcl6, an inhibitor of ASC differentiation, ectopic Zbtb20 expression in primary B cells facilitates terminal B cell differentiation to ASCs. In plasma cell lines, Zbtb20 induces cell survival and blocks cell cycle progression. Immunized Zbtb20-deficient mice exhibit curtailed humoral responses and accelerated loss of antigen-specific plasma cells, specifically from the BM pool. Strikingly, Zbtb20 induction does not require Blimp1 but depends directly on Irf4, acting at a newly identified Zbtb20 promoter in ASCs. These results identify Zbtb20 as an important player in late B cell differentiation and provide new insights into this complex process.

Mice lacking a functional gene for the Oct2 transcriptional activator display several developmental and functional deficiencies in the B lymphocyte lineage. These include defective B cell receptor (BCR) and Toll-like receptor 4 signaling, an absence of B-1 and marginal zone populations, and globally reduced levels of serum immunoglobulin (Ig) in naive and immunized animals. Oct2 was originally identified through its ability to bind to regulatory regions in the Ig loci, but genetic evidence has not supported an essential role for Oct2 in the expression of Ig genes. We describe a new Oct2-mediated role in B cells. Oct2 augments the ability of activated B cells to differentiate to antibody-secreting plasma cells (ASCs) under T cell–dependent conditions through direct regulation of the gene encoding the α chain of the interleukin (IL) 5 receptor. Ectopic expression of IL-5Rα in oct2-deficient B cells largely restores their ability to differentiate to functional ASCs in vitro but does not correct other phenotypic defects in the mutants, such as the maturation and specialization of peripheral B cells, which must therefore rely on distinct Oct2 target genes. IL-5 augments ASC differentiation in vitro, and we show that IL-5 directly activates the plasma cell differentiation program by enhancing blimp1 expression.

Normalization is critical for removing systematic variation from microarray data. For two-color microarray platforms, intensity-dependent lowess normalization is commonly used to correct relative gene expression values for biases. Here we outline a normalization method for use when the assumptions of lowess normalization fail. Specifically, this can occur when specialized boutique arrays are constructed that contain a subset of genes selected to test particular biological functions.

Resting B cells can be cultured to induce antibody-secreting cell (ASC) differentiation in vitro. A quantitative analysis of cell behavior during such a culture allows the influences of different stimuli and gene products to be measured. The application of this analytical system revealed that the OBF-1 transcriptional coactivator, whose loss impairs antibody production in vivo, has two effects on ASC development. Although OBF-1 represses early T cell–dependent (TD) differentiation, it is also critical for the completion of the final stages of ASC development. Under these conditions, the loss of OBF-1 blocks the genetic program of ASC differentiation so that Blimp-1/prdm1 induction fails, and bcl-6, Pax5, and AID are not repressed as in control ASC. Retroviral complementation confirmed that OBF-1 was the critical entity. Surprisingly, when cells were cultured in lipopolysaccharide to mimic T cell–independent conditions, OBF-1–null B cells differentiated normally to ASC. In the OBF-1^−/− ASC generated under either culture regimen, antibody production was normal or only modestly reduced, revealing that Ig genes are not directly dependent on OBF-1 for their expression. The differential requirement for OBF-1 in TD ASC generation was confirmed in vivo. These studies define a new regulatory role for OBF-1 in determining the cell-autonomous capacity of B cells to undergo terminal differentiation in response to different immunological signals.