ReviewThe genetic network controlling plasma cell differentiation
Highlights
► Terminal differentiation of B cells to plasma cells occurs in a stochastic manner. ► A gene regulatory network controls plasma cell differentiation. ► Gene expression in B cells and plasma cells is maintained by distinct transcription factors. ► Blimp1 and Irf4 are dose-dependent regulators of plasma cells.
Introduction
Terminal differentiation is a generally irreversible process that, in the haematopoietic system, leads to the acquisition of specialized effector functions and exit from the cell cycle. The terminal differentiation process is also associated with profound alterations in the morphology, lifespan and gene expression profiles of the differentiated cells compared to their predecessors. The transition of B cells into antibody-secreting cells (ASCs) represents one such terminal differentiation process that is essential for the adaptive immune response.
There are three main subsets of mature B cells; follicular B cells, which represent the majority of naïve B cells, marginal zone (MZ) and B1 B cells. MZ B cells reside in the marginal sinus of the spleen, whereas B1 B cells occur predominantly in the peritoneal cavity and at mucosal sites [1]. In contrast, conventional or follicular B cells are almost exclusively found in the lymphoid follicles of the spleen and lymph nodes. The relative propensity of B cells to undergo terminal differentiation varies between the subsets, with MZ and B1 B cells being specialized, both geographically and genetically, for rapid response to so-called T-independent antigens (TI), such as bacterial components [2]. Follicular B cells may also respond to TI antigens, but appear specialized for responding to antigens that also elicit CD4+ helper T cell responses. Upon antigen encounter and receipt of T cell help, follicular B cells undergo multiple rounds of division and have the unique ability to differentiate into both ASCs (plasmablasts and plasma cells) and memory B cells [1]. Plasmablasts are short-lived, cycling ASCs that are found in extrafollicular foci in peripheral lymphoid organs. Plasmablasts have often undergone immunoglobulin class switch recombination (CSR), but not somatic hypermutation (SHM) to increase the affinity of the resulting antibodies for the antigen. In a poorly understood process, some activated B cells return to the B cell follicle and proliferate vigorously to form a germinal centre (GC), where the B cells initiate SHM to enable further clonal selection. The GC produces long-lived plasma cells that are non-cycling and home preferentially to the bone marrow [1]. The GC reaction also produces memory B cells. These cells maintain a B cell phenotype but can rapidly differentiate into ASCs following re-exposure to antigen.
The process of B cell terminal differentiation can be studied in vitro, as B cells are capable of both CSR and ASC differentiation in response to T-cell derived stimuli (CD40 ligation and cytokines) or TI-related signals (Toll-like receptor (TLR) ligation by pathogen derived products such as lipopolysaccharide (LPS) and hyper-methylated CpG DNA). B cell responses in vitro thus provide a controlled system to investigate the biology of B cell terminal differentiation on both a cellular and molecular level.
Quantitative analysis of in vitro B cell cultures has revealed a striking relationship between cell division history and CSR and ASC differentiation [3], [4], [5]. The proportion of B cells that undergo either of these differentiation events typically increases with each consecutive division (Fig. 1). The time taken to traverse each cell cycle is highly variable between cells, but does not seem to alter the differentiation rate, implying that cell division itself is playing a critical role in tuning the molecular machinery controlling CSR and the generation of ASCs. Furthermore, T cell cytokines such as IL4 and IL5 alter the probability of CSR and ASC differentiation with division (Fig. 1B). Hasbold et al. found that CSR and development of ASCs behaved as independent stochastic processes, allowing the numbers of class switched ASCs in each division to be predicted for different cytokine concentrations and combinations [5]. These findings have lead to a division-based model of B cell behaviour that describes how stochastic decisions taken at a single cell level result in the controlled generation of a variety of differentiated cell types in the population as a whole [5].
Complementing these cellular studies, the last few years have seen major advances in our understanding of the transcriptional regulation of the B cell to ASC transition [6], [7]. The gene expression changes that are required for this process are regulated by the coordinated activity of a small group of so-called master regulatory transcription factors. These factors can be divided into those, such as Pax5 and Bcl6, which promote and maintain the B cell program, and others such as Blimp1 and Irf4 that control ASC differentiation. The B cell and ASC factors appear to regulate mutually antagonistic transcriptional programs resulting in a gene regulatory network that ensures the separation of the B cell and ASC fates [6].
While most studies to date have proposed that these master regulatory transcription factors function in a dominant manner to either maintain the B cell fate or to drive ASC differentiation, such deterministic behaviour has not yet been reconciled with the evidence supporting stochastic, division-based regulation of differentiation outlined above. In this review we will discuss both the transcriptional and cellular models of B cell terminal differentiation to draw attention to the need for a model of the genetic network of ASC differentiation that more accurately describes the flexible, dynamic and complex regulation of the adaptive immune response.
Section snippets
Factors that promote the B cell fate
One class of transcription factors can be conveniently grouped into those whose primary function is to promote the B cell gene expression program and to prevent ASC differentiation. Here we have focused on the best-characterized B cell factors: Pax5, Bcl6, Bach2 and Oct2/Obf1. It should be noted, however, that a number of other transcriptional regulators, including PU.1 [8], Irf8 [9], MITF [10] and Ets1 [11] are also thought to play roles in the process (Fig. 2).
Irf4
Irf4 is a multi-functional transcriptional regulator that controls many aspects of B cell differentiation including Igκ gene recombination, CSR, GC B cell formation and ASC differentiation [47]. Irf4 also is broadly required for the differentiation of CD4+ T cells [48]. Irf4 can bind to DNA weakly on its own, but displays strong co-operative binding in the presence of PU.1, or the closely related Spi-B [49], [50]. Irf4-PU.1 dimers have been shown to be important in the regulation of the Igh and
Towards describing a gene regulatory network controlling late B cell differentiation
The large amount of data that was summarized in the preceding section was in the whole generated by the analysis of individual transcription factors in isolation. However, this information can also be used to construct gene regulatory networks that attempt to define the crucial processes and interactions that allow the transition from a B cell to an ASC, in a manner analogous to that successfully applied to early haematopoiesis and T cell differentiation [80], [81]. B cell terminal
Blimp1 and Irf4 are dose-dependent regulators of antibody-secreting cell differentiation
While Irf4 emerges as a key component in the decision to undergo CSR and develop into an ASC, it is necessary to contrast its action with the patterned division-linked changes seen at the population level. Questions to be addressed include: why do only a proportion of stimulated cells in vitro develop into ASCs and undergo CSR in each division, despite the presence of Irf4 in all activated B cells? If Irf4 plays a critical role for both CSR and ASC development, why are the two outcomes poorly
Conclusions and future perspectives
Research over the past decade has provided a wealth of data to highlight the key factors in the gene regulatory network driving the terminal differentiation of B cells. Two classes of transcription factors are required, those that promote the B cell state, such as Bcl6 and Pax5 and those including Irf4 and Blimp1, that favour ASC differentiation. On a genetic level, the mutual antagonism between these factors ensures that these key developmental stages in B cell differentiation are kept
Acknowledgements
We would like to thank members of the Walter and Eliza Hall Institute B cell program for discussions. This research was supported by the Pfizer Australia Research Fellowship and ARC Future Fellowship to S.L.N. and National Health and Medical Research Council of Australia Research Fellowships to P.D.H. and L.M.C. This work was made possible through Victorian State Government Operational Infrastructure Support and Australian Government NHMRC IRIIS.
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2022, Cell ReportsCitation Excerpt :Of note, prdm1 RNA (encoding BLIMP1) was lower in DNT-B cells as well (Figure 4J). To exlude the possibility that BLIMP1 was repressed by high abundance of BCL6 (Nutt et al., 2011) we also examined BCL6 protein (Figures 4H and 4I), showing that diminished BLIMP1 was not due to elevated BCL6. To corroborate these findings in vivo and to exclude potentially confounding effects of non-mutant B cells, we labeled 48-h LPS-activated B cells with a proliferation dye and transferred them into rag2−/− mice (Figure S4C).