Development of B cells expressing surface immunoglobulin molecules that lack V(D)J-encoded determinants in the avian embryo bursa of Fabricius

Abstract

Immunoglobulin gene rearrangement in avian B cell precursors generates surface Ig receptors of limited diversity. It has been proposed that specificities encoded by these receptors play a critical role in B lineage development by recognizing endogenous ligands within the bursa of Fabricius. To address this issue directly we have introduced a truncated surface IgM, lacking variable region domains, into developing B precursors by retroviral gene transfer in vivo. Cells expressing this truncated receptor lack endogenous surface IgM, and the low level of endogenous Ig rearrangements that have occurred within this population of cells has not been selected for having a productive reading frame. Such cells proliferate rapidly within bursal epithelial buds of normal morphology. In addition, despite reduced levels of endogenous light chain rearrangement, those light chain rearrangements that have occurred have undergone variable region diversification by gene conversion. Therefore, although surface expression of an Ig receptor is required for bursal colonization and the induction of gene conversion, the specificity encoded by the prediversified receptor is irrelevant and, consequently, there is no obligate ligand for V(D)Jencoded determinants of prediversified avian cell surface IgM receptor.The rearrangement of chicken Ig genes generates minimal antibody diversity. At the light chain (L) locus, the unique, functional V L 1 gene rearranges to the unique J L segment in all B cells (1). Similarly, at the heavy chain (H) locus, unique V H 1 and J H genes undergo rearrangement with a family of highly conserved D H elements to form a VDJ H complex of limited diversity (2, 3). Junctional diversity is limited further because of the lack of N nucleotide additions in chicken V(D)J junctions (4). A diverse repertoire of B cell specificities is generated by somatic gene-conversion events in which VD H and V L sequences in functionally rearranged VDJ H and VJ L genes are replaced by homologous sequences from upstream V L and V H pseudogenes (⌿V L and ⌿V H ) (2, 5-7).The bursa of Fabricius is the primary site of B cell lymphopoiesis in avian species, and surgical or chemical ablation of the chicken bursa profoundly disrupts the normal progression of B cell development and the generation of diversity by gene conversion (reviewed in refs. 7-10). The bursa is colonized by a single wave of B cell precursors during embryogenesis starting at about embryonic day 8 (E8) and lasting about a week (11). Within the bursa, B cell precursors first are observed in the mesenchymal tissue, and 20,000-40,000 precursors subsequently migrate across the bursal epithelial basement membrane and begin to proliferate in oligoclonal clusters or epithelial buds from which the discrete follicles of the mature bursa are derived (12, 13).Surface Ig (sIg) ϩ B cell precursors first are observed by about E9-E10, and the frequency of such cells increases rapidly during the embryonic period (14). Although Ig gene rearrangement is not intrinsically biased towar...

The sheep and cattle Peyer’s patch as a site of B-cell development

“…After birth, antigen stimulation via bursal epithelium plays a role in the selection of B-cell clones [9]. Using retroviral gene transfer techniques which induce truncated IgM as B-cell receptor, it is clearly demonstrated that the regulation of chicken bursal B-cell development after hatching differs from that in the embryo [107,108]. Therefore further studies are required to determine the effect of external antigens on the selection of B-cells within follicles of both types of PP follicles as well as the functional difference between the ileal and jejunal PP.…”

Role of Commensal Bacteria in Development of Gut-Associated Lymphoid Tissues and Preimmune Antibody Repertoire

“…Transgenic chickens whose B cells expressed a truncated (V H -less) form of surface IgM underwent rapid cell death in the bursa after hatching, suggesting that signaling through this receptor, presumably by lumenal Ags, is required for proliferation and/or survival of B cells (44).…”

Dual Requirement for the Igα Immunoreceptor Tyrosine-Based Activation Motif (ITAM) and a Conserved Non-Igα ITAM Tyrosine in Supporting Igαβ-Mediated B Cell Development

“…Line O chicken embryo fibroblasts (CEFs; Regional Poultry Research Laboratories) were transfected with the RCAS-mCD8:chIg vectors as previously described (13). One week after transfection, CEFs were tested for surface expression of the mCD8:chIg chimeric proteins and injected into line 22 chick embryos (Charles River Laboratories) on day 3 of embryogenesis.…”

“…Cell suspensions of bursal cells generated at the time of hatching were also stained for the pan B cell marker ChB6 and as previously described (13).…”

“…It has been suggested that such basal signaling, initiated from the BCR complex in the absence of receptor ligation, is sufficient to support the development of B cells. To address this possibility, both murine and avian truncated receptor complexes (T), which lack the V(D)J encoded determinants, but maintain the ability to associate with the Ig␣␤ heterodimer, have been generated and have been shown to support early B cell development (13)(14)(15). Although these results suggested that there is no requirement for BCR ligation during the early stages of B cell development in mouse or in chicken, both murine and avian T complexes included extracellular domains that could have been involved in receptor-ligand interactions.…”