Biomedical subjects

Michael J H Ratcliffe

Publications and source records attributed to Michael J H Ratcliffe.

9 recordsLinked to original sources

PubMed2006

Antibodies, immunoglobulin genes and the bursa of Fabricius in chicken B cell development.

The bursa of Fabricius is critical for the normal development of B lymphocytes in birds. It is productively colonized during embryonic life by a limited number of B cell precursors that have undergone the immunoglobulin gene rearrangements required for expression of cell surface immunoglobulin. Immunoglobulin gene rearrangement occurs in the absence of terminal deoxynucleotidyl transferase and generates minimal antibody diversity. In addition, observations that immunoglobulin heavy and light chain variable gene rearrangement occur at the same time and that allelic exclusion of immunoglobulin expression is regulated at the level of variable region gene rearrangement provide a striking contrast to rodent and primate models of immunoglobulin gene assembly. Following productive colonization of the bursa, developing B cells undergo rapid proliferation and the immunoglobulin V region genes that generate the specificity of the B cell surface immunoglobulin receptor undergo diversification. Immunoglobulin diversity in birds is generated by somatic gene conversion events in which sequences derived from upstream families of pseudogenes replace homologous sequences in unique and functionally rearranged immunoglobulin heavy and light chain variable region genes. This mechanism is distinct from and much more efficient than mechanisms of antibody diversification seen in rodents and primates. While the bursal microenvironment is not required for immunoglobulin gene rearrangement and expression, it is essential for the generation of antibody diversity by gene conversion. Following hatch, gut derived antigens are taken up by the bursa. While bursal development prior to hatch occurs in the absence of exogenous antigen, chicken B cell development after hatch may therefore be influenced by the presence of environmental antigen. This review focuses on the differences between B cell development in the chicken as compared to rodent and primate models.

Animals↗

PubMed2006

Ligand-independent signaling during early avian B cell development.

Surface immunoglobulin (sIg) expression has been conserved as a critical checkpoint in B lymphocyte development. In the chicken embryo, only sIg+ B cells are selectively expanded in the bursa of Fabricius, a primary lymphoid organ unique to the avian species. We have previously demonstrated that an interaction between the antigen- binding sites of sIg and a specific bursal ligand(s) is not required to regulate this developmental checkpoint. Rather, the requirement for sIg expression can be attributed to the surface expression of the Igalpha/beta heterodimer associated with sIg. More specifically, ligand-independent signaling downstream of the Igalpha cytoplasmic domain drives all bursal stages of B cell development during embryogenesis. We discuss here a site-directed mutagenesis approach to identify the critical membrane proximal events involved in ligand-independent signaling during B cell development.

Animals↗

PubMed2005

Cell surface immunoglobulin regulated checkpoints in chicken B cell development.

The bursa of Fabricius is critical for the normal development of B lymphocytes in avian species. Productive colonization of bursal follicles by B cell precursors requires surface immunoglobulin expression. We have shown using retroviral gene transfer that expression of chimeric receptors containing the extracellular and transmembrane domains of murine CD8alpha and CD8beta fused to the cytoplasmic domains of chicken Igalpha and Igbeta can support productive bursal colonization in the chicken embryo in bursal cells lacking the expression of endogenous sIgM. We show here that chimeric receptor expression does not support continued bursal cell development after hatch. However intrabursal administration of anti-CD8 antibodies that ligate the CD8alpha:Igalpha chimeric receptor results in maintained numbers of bursal cells that express the chimeric receptor in the absence of endogenous sIgM. These results support a model in which sIgM receptor expression is required for productive bursal colonization in the chick embryo but sIgM receptor ligation is required to support later B cell development after hatch.

Animals↗

PubMed2005

Dual requirement for the Ig alpha immunoreceptor tyrosine-based activation motif (ITAM) and a conserved non-Ig alpha ITAM tyrosine in supporting Ig alpha beta-mediated B cell development.

Surface Ig (sIg) expression is a critical checkpoint during avian B cell development. Only cells that express sIg colonize bursal follicles, clonally expand, and undergo Ig diversification by gene conversion. Expression of a heterodimer, in which the extracellular and transmembrane domains of murine CD8alpha or CD8beta are fused to the cytoplasmic domains of chicken Igalpha (chIgalpha) or Igbeta, respectively (murine CD8alpha (mCD8alpha):chIgalpha + mCD8beta:chIgbeta), or an mCD8alpha:chIgalpha homodimer supported bursal B cell development as efficiently as endogenous sIg. In this study we demonstrate that B cell development, in the absence of chIgbeta, requires both the Igalpha ITAM and a conserved non-ITAM Igalpha tyrosine (Y3) that has been associated with binding to B cell linker protein (BLNK). When associated with the cytoplasmic domain of Igbeta, the Igalpha ITAM is not required for the induction of strong calcium mobilization or BLNK phosphorylation, but is still necessary to support B cell development. In contrast, mutation of the Igalpha Y3 severely compromised calcium mobilization when expressed as either a homodimer or a heterodimer with the cytoplasmic domain of Igbeta. However, coexpression of the cytoplasmic domain of Igbeta partially complemented the Igalpha Y3 mutation, rescuing higher levels of BLNK phosphorylation and, more strikingly, supporting B cell development.

Adaptor Proteins, Signal Transducing↗

PubMed2004

The cytoplasmic domain of Ig alpha is necessary and sufficient to support efficient early B cell development.

The B cell receptor complex (BcR) is essential for normal B lymphocyte function, and surface BcR expression is a crucial checkpoint in B cell development. However, functional requirements for chains of the BcR during development remain controversial. We have used retroviral gene transfer to introduce components of the BcR into chicken B cell precursors during embryonic development. A chimeric heterodimer, in which the cytoplasmic domains of chicken Igalpha and Igbeta are expressed by fusion with the extracellular and transmembrane domains of murine CD8alpha and CD8beta, respectively, targeted the cytoplasmic domains of the BcR to the cell surface in the absence of extracellular BcR domains. Expression of this chimeric heterodimer supported all early stages of embryo B cell development: bursal colonization, clonal expansion, and induction of repertoire diversification by gene conversion. Expression of the cytoplasmic domain of Igalpha, in the absence of the cytoplasmic domain of Igbeta, was not only necessary, but sufficient to support B cell development as efficiently as the endogenous BcR. In contrast, expression of the cytoplasmic domain of Igbeta in the absence of the cytoplasmic domain of Igalpha failed to support B cell development. The ability of the cytoplasmic domain of Igalpha to support early B cell development required a functional Igalpha immunoreceptor tyrosine-based activation motif. These results support a model in which expression of surface IgM following productive V(D)J recombination in developing B cell precursors serves to chaperone the cytoplasmic domain of Igalpha to the B cell surface, thereby initiating subsequent stages of development.

Amino Acid Motifs↗

PubMed2004

The avian B-cell receptor complex: distinct roles of Igalpha and Igbeta in B-cell development.

The bursa of Fabricius has evolved in birds as a gut-associated site of B-cell lymphopoiesis that is segregated from the development of other hematopoietic lineages. Despite differences in the developmental progression of chicken as compared to murine B-cell lymphopoiesis, cell-surface immunoglobulin (sIg) expression has been conserved in birds as an essential checkpoint in B-cell development. B-cell precursors that express an sIg complex that includes the evolutionarily conserved Igalpha/beta heterodimer colonize lymphoid follicles in the bursa, whereas B-cell precursors that fail to express sIg due to non-productive V(D)J recombination are eliminated. Productive retroviral gene transfer has allowed us to introduce chimeric receptor constructs into developing B-cell precursors in vivo. Chimeric proteins comprising the extracellular and transmembrane regions of murine CD8alpha fused to the cytoplasmic domain of chicken Igalpha efficiently supported B-cell development in precursors that lacked endogenous sIg expression. By contrast, expression of an equivalent chimeric receptor containing the cytoplasmic domain of Igbeta actively inhibited B-cell development. Consequently, the cytoplasmic domains of Igalpha and Igbeta play functionally distinct roles in chicken B-cell development.

Amino Acid Sequence↗

PubMed2002

B cell development in gut associated lymphoid tissues.

B lymphocyte development can occur in a variety of anatomical sites. While typically considered to be a process that occurs in the bone marrow throughout life, it is becoming clear that gut associates sites of B cell development are critically important in many species of veterinary importance. Among these sites, the bursa of Fabricius in chickens and the ileal Peyer's patches of sheep are among the best studied. In these organs, it has become clear that many of the properties associated with B cell development in rodent and primate bone marrow do not apply. Thus while bone marrow B cell development typically involves an ongoing maturation of mature B cells from immature B lineage precursors that lack the expression of a surface immunoglobulin complex, gut associated lymphoid tissues (GALTs) may be colonized by a single wave of precursor cells during embryo development. Nonetheless, molecular analysis of the requirements for B lymphocyte development in GALTs reveals some striking parallels with requirements identified for B cell development in bone marrow. This article will discuss differences between B cell development in the bone marrow and GALTs and recent evidence emerging that yields insights into how these processes are regulated.

Animals↗

PubMed2002

Influence of antibody diversification on the mechanism of haplotype exclusion of immunoglobulin gene expression.

Allelic, or haplotype, exclusion of immunoglobulin gene expression ensures that the products of a single allele or light chain isotype are expressed on the B cell surface. Evidence has accumulated in rodent and primate models to indicate that the products of successful rearrangement regulate this process. In contrast, haplotype exclusion of chicken immunoglobulin gene expression is regulated at the level of variable region gene rearrangement. We discuss here alternative models for ensuring haplotype exclusion that may operate in the chicken and extend the discussion to address the issue as to how two apparently distinct mechanisms may have evolved to yield the same outcome.

Alleles↗

PubMed2002

Cell surface immunoglobulin receptors in B cell development.

Expression of surface immunoglobulin (sIg) related receptors has been conserved in phylogenetically distinct species as a critical checkpoint in B cell development. The sIg receptor comprises extracellular IgM heavy and light chains, with the potential for ligand binding, complexed to the Igalpha/Igbeta heterodimer that is responsible for signal transduction through sIg. Experimental systems, from both avian and murine models of B cell development, have been designed to identify the function of individual receptor components in B cell development. In this review, we assess the regulatory functions of different components of the sIg receptor complex during early development in experimental systems from evolutionarily distinct species.

Animals↗