In and Out of the Bursa—The Role of CXCR4 in Chicken B Cell Development

Abstract

In contrast to mammals, early B cell differentiation and diversification of the antibody repertoire in chickens do not take place in the bone marrow but in a specialized gut associated lymphoid tissue (GALT), the bursa of Fabricius. During embryonic development, B cell precursors migrate to the bursa anlage, where they proliferate and diversify their B cell receptor repertoire. Around hatch these diversified B cells start to emigrate from the bursa of Fabricius and populate peripheral lymphoid organs, but very little is known how the migratory processes are regulated. As CXCL12 (syn. SDF-1) and CXCR4 were shown to be essential for the control of B cell migration during the development of lymphoid tissues in mammals, we analyzed expression and function of this chemokine/chemokine-receptor pair in the chicken bursa. We found a strong variation of mRNA abundance of CXCL12 and CXCR4 in different stages of bursa development, with high abundance of CXCL12 mRNA in the bursa anlage at embryonic day 10 (ED10). In situ hybridization demonstrated disseminated CXCL12 expression in the early bursa anlage, which condensed in the developing follicles and was mainly restricted to the follicle cortex post-hatch. Flow cytometric analysis detected CXCR4 protein already on early B cell stages, increasing during bursal development. Post-hatch, a subpopulation with the hallmarks of emigrating B cells became detectable, which had lower CXCR4 expression, suggesting that downregulation of CXCR4 is necessary to leave the CXCL12-high bursal environment. In vivo blockade of CXCR4 using AMD3100 at the time of B cell precursor immigration strongly inhibited follicle development, demonstrating that CXCL12 attracts pre-bursal B cells into the bursal anlage. Altogether, we show that CXCL12 and its receptor CXCR4 are important for both populating the bursa with B cells and emigration of mature B cells into the periphery post hatch, and that CXCR4 function in primary B cell organs is conserved between mammals and birds.

Full-Length Transcriptome: A Reliable Alternative for Single-Cell RNA-Seq Analysis in the Spleen of Teleost Without Reference Genome

“…Compared with mammals, the early B cell differentiation and diversification of antibody pools in birds do not occur in the bone marrow, but in a special gut related lymphoid tissue, the bursa of Fabricius ( 84 ). During embryonic development, B cell precursors migrate to the bursa primordium, where they proliferate and diversify the B cell receptor pool ( 85 ).…”

“…Around hatch, these diverse B cells begin to migrate from the bursa of Fabricius to the peripheral lymphoid organs, but little is known about the regulation of the migration process (86). Masteller et al proved that changes in cell surface glycosylation may be related to the colonization of the bursa of Fabricius (84). Prebursal and early B cells express the carbohydrate epitope sialyl Lewis(x) (CD15s), which is a carbohydrate moiety that participates in the adhesion of leukocytes to endothelial cells through selectin binding (84).…”

“…Masteller et al proved that changes in cell surface glycosylation may be related to the colonization of the bursa of Fabricius (84). Prebursal and early B cells express the carbohydrate epitope sialyl Lewis(x) (CD15s), which is a carbohydrate moiety that participates in the adhesion of leukocytes to endothelial cells through selectin binding (84). In contrast, the B cells in the bursa of Fabricius were transformed into high-level expression of Lewis (x) (CD15) (87) after gene conversion.…”

The Chicken Embryo Model: A Novel and Relevant Model for Immune-Based Studies

“…Because they only have a single copy of functional V and J segments for both chains, the chicken immune system relies on another mechanism known as somatic gene conversion, a process taking place during bursal development (134,136). Thus, the clusters of pseudogenes, upstream of the immunoglobulin loci, are critical for antibody diversity in chickens (139,140). After a low-efficiency V(D)J rearrangement in the bone marrow, immature B cells migrate to the bursa of Fabricius where gene conversation, a process in which V sequences are replaced with the pseudogene sequences, takes place.…”

“…After maturation and TCR rearrangement, gd T cells migrate to the spleen and intestinal epithelium by ED15, whereas ab T cells only start doing so at ED19 (27,133,194,195). Meanwhile, the bursa of Fabricius rudiment is colonized by lymphoid progenitors between ED8 and ED14 (136,140,186). B cells can first be detected there at ED11-12, and like T cells, are believed to be fully immunocompetent by ED18 (14,193,196).…”