For this study we have used

For this study we have used the intermediate affinity mutant antigen HEL2× for immunization. However, we have observed a similar defect in the plasmablast differentiation of EBI2-deficient SWHEL vegf pathway over a 10,000-fold affinity range by using WT HEL or the low-affinity mutant HEL3× (data not shown). This indicates that increasing or decreasing BCR signal strength, which is known to regulate the plasmablast response (Benson et al., 2007, Paus et al., 2006), does not correct or exacerbate the defective response of EBI2-deficient B cells. Consistent with this result, the in vitro activation and proliferation of B cells was not affected by absence of EBI2, and therefore a contribution of EBI2 to the signals required for efficient B cell stimulation is unlikely. Notably, expression of EBI2 was also not required for normal in vitro plasmablast differentiation. It is still possible, however, that signals delivered by EBI2 can directly trigger or regulate gene expression programs that drive plasmablast differentiation. On the other hand, the guidance of responding B cells to distinct microenvironments mediated by modulation of EBI2 expression may subject them to alternative extracellular milieus that could direct their subsequent lineage commitment. Thus, EBI2-deficient B cells may predominantly form GC B cells in vivo because of the fact that they remain in a microenvironment proximal to the FDC network, in which antigen deposits and follicular T helper cells drive their proliferation and selection into the long-term effectors of humoral immunity. At the same time, because the factors and myeloid cell populations supporting maturation and survival of plasmablasts are concentrated outside of B cell follicles, they are not encountered by most responding EBI2-deficient B cells (Garcia De Vinuesa et al., 1999, Mohr et al., 2009). The control of EBI2 expression, via NF-κB, Bcl-6, or other pathways, therefore plays an important role in determining the balance between plasmablast and GC differentiation. In vitro, cytokines such as interleukin (IL)-4, IL-6, and IL-10 have been shown to modulate Gpr183 expression (Lam et al., 2008, Shaffer et al., 2000). In contrast, we have only seen minimal differences in the expression of Gpr183 mRNA in SWHEL B cells stimulated with HEL antigen of different affinities (data not shown). Although regulation of EBI2 is an important component of the early commitment of responding B cells to the extrafollicular versus GC pathway of differentiation, it is clear that this fundamental decision in B cell fate is a complex interplay of many signals in vivo. However, it remains possible that abnormal regulation of EBI2 expression leads to autoimmune and inflammatory diseases in which the balance between plasmablast and GC B cell differentiation is lost. The importance of EBI2 for B cell function was first suggested by the dramatic upregulation of this receptor in EBV-transformed B cells and further inferred from its regulation in activated and GC B cells (Birkenbach et al., 1993, Glynne et al., 2000, Shaffer et al., 2000). An involvement of EBI2 in pathology has also been suggested by its dysregulated expression in B cell-associated autoimmune and neoplastic diseases (Aalto et al., 2001, Ye et al., 2003). However, the function of EBI2 and the significance and implications of its modulation have long awaited clarification. This study provides evidence for a biological function of EBI2 and indicates that this receptor provides an extra dimension to B cell migration and differentiation. Modulation of EBI2 expression is necessary for ensuring both the rapid and long-term antibody production that are required for optimal protection against pathogens. Identification of the putative ligand for EBI2 and elucidation of the molecular mechanisms by which it controls B cell migration and differentiation may prove valuable in designing new vaccine strategies and potential therapeutics for immune disorders.