br STAR Methods br Microenvironmental niches for na
Microenvironmental niches for naïve and activated B cells Secondary lymphoid organs, such as the spleen and lymph nodes, are structurally and functionally compartmentalized to provide a set of microenvironmental niches that support the survival of resting B cells, as well as the activation and differentiation of 52 6 participating in antibody responses. In secondary lymphoid organs, B and T cells segregate to form B cell follicles that surround a central T cell zone (Figure 1). In the spleen, B cell follicles are surrounded by the marginal zone, an area rich in macrophages that also harbors a population of resident marginal zone B cells. Dendritic cell (DC) populations are localized in T cell areas as well as in interfollicular areas and the splenic marginal zone bridging channels, which are breaks in the marginal sinus where the T cell zone abuts the red pulp. Central to the compartmentalization of lymphoid organs is the localized production of chemokines and other guiding cues by networks of stromal cells (Table 1). The chemokine CXC ligand (CXCL)13 is made by follicular stromal cells, which comprise the marginal reticular cells in the outer follicle and a central network of follicular dendritic cells (FDCs) . Naïve recirculating B cells respond primarily to this chemokine as a result of their high expression of chemokine CXC receptor (CXCR)5 and therefore migrate to B cell follicles . By contrast, the fibroblastic reticular cell network in T cell zones is characterized by expression of the chemokines CC ligand (CCL)19 and CCL21, which mediate the migration of chemokine CC receptor (CCR)7-expressing T cells to these areas . During antibody responses, B cell migration is orchestrated to facilitate: (i) access to antigen; (ii) interactions with helper T cells; and (iii) homing to the specialized microenvironments that promote rapid or long-term antibody production. Naïve B cells patrol secondary lymphoid organs for the presence of antigen by rapid, random movement within follicles. After antigen encounter, B cells move to the boundary between B cell follicles and T cell areas where they interact with T helper cells and undergo initial proliferation (Figure 2) . This movement occurs about 6h after activation and is directed by the antigen-mediated upregulation of CCR7; the receptor for the T cell zone chemokines CCL19 and CCL21 . Early in the response, proliferating B cells begin to follow one of two alternate fates by differentiating into short-lived plasma cells or germinal center (GC) B cells. B cells recruited into the former pathway migrate to extrafollicular sites (Figure 2) where their proliferation and differentiation into plasmablasts and plasma cells is sustained . Alternatively, B cells can localize to the FDC-dense areas in the center of follicles where they seed GCs (Figure 2) 7, 8. As the response progresses, B cells leave GC reactions either as long-lived plasma cells, which take up residency in the bone marrow, or as memory B cells . It has only recently been appreciated that antigen-activated B cells undertake two additional transient migrations during the early stages of T-dependent antibody responses. In the first of these, antigen-activated B cells migrate rapidly (within 2–3h) to the outer regions of the follicle before they relocate to the T–B boundary to receive T cell help (Figure 2) . After interacting with T cells at the T–B boundary, activated B cells have also been shown to relocalize to interfollicular and outer follicular regions prior to their differentiation into early plasmablasts or entry into GCs (Figure 2) 10, 11, 12, 13. It transpires that both of these early B cell migratory steps are mediated by the chemoattractant receptor EBI2 (also known as GPR183) and its oxysterol ligand 9, 14, 15, 16, 17. In addition, modulation of chemotactic signal delivered to responding B cells through EBI2 plays a critical role in delivering them to the correct microenvironments to undergo plasmablast versus GC B cell differentiation. Thus, EBI2 is part of a network of chemoattractant receptors that regulates B cell localization, providing an extra dimension to the migratory cues provided by conventional chemokines and their receptors. Here, we review the role of EBI2 and oxysterols in controlling B cell migration and discuss how EBI2 collaborates with the chemokine receptors CXCR5, CCR7, and CXCR4 to position B cells appropriately during T-dependent antibody responses.