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

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.

Identification of the Receptor and Cellular Ortholog of the Marek's Disease Virus (MDV) CXC Chemokine.

Marek's disease virus (MDV) is a cell associated alphaherpesvirus that causes fatal lymphoma in chickens. One factor that plays a crucial role in MDV pathogenesis is the viral CXC chemokine vIL-8 that was originally named after chicken interleukin 8 (cIL-8). However, a recent study demonstrated that vIL-8 recruits B cells and a subset of T cells but not neutrophils, suggesting that vIL-8 is not a cIL-8 orthologue. In this study, we set to identify the cellular orthologues and receptor of vIL-8 using in silico analyses, binding and chemotaxis assays. Sequence and phylogenetic analyses of all chicken CXC chemokines present in the recently published chicken genome revealed that vIL-8 shares the highest amino acid similarity with the CXCL13L1 variant. To evaluate if vIL-8 and CXCL13L1 are also functional orthologues, we assessed their binding properties and chemotaxis activity. We demonstrated that both vIL-8 and CXCL13 variants bind B cells and subsets of T cells, confirming that they target the same cell types. In addition, the chemokines not only bound the target cells but also induced chemotaxis. Finally, we identified CXCR5 as the receptor of vIL-8 and CXCL13 variants and confirmed that the receptor is expressed on MDV target cells. Taken together, our data demonstrate the conservation of the receptor-ligand interaction between CXCR5 and CXCL13 and shed light on the origin and function of the MDV-encoded vIL-8 chemokine, which plays a crucial role in the pathogenesis of this highly oncogenic virus.