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J Donald Capra

Publications and source records attributed to J Donald Capra.

10 recordsLinked to original sources
PubMed2006

DNA cleavage of a cryptic recombination signal sequence by RAG1 and RAG2. Implications for partial V(H) gene replacement.

Antibody and T cell receptor genes are assembled from gene segments by V(D)J recombination to produce an almost infinitely diverse repertoire of antigen specificities. Recombination is initiated by cleavage of conserved recombination signal sequences (RSS) by RAG1 and RAG2 during lymphocyte development. Recent evidence demonstrates that recombination can occur at noncanonical RSS sites within Ig genes or at other loci, outside the context of normal lymphocyte receptor gene rearrangement. We have characterized the ability of the RAG proteins to bind and cleave a cryptic RSS (cRSS) located within an Ig V(H) gene segment. The RAG proteins bound with sequence specificity to either the consensus RSS or the cRSS. The RAG proteins nick the cRSS on both the top and bottom strands, thereby bypassing the formation of the DNA hairpin intermediate observed in RAG cleavage of canonical RSS substrates. We propose that the RAG proteins may utilize an alternative mechanism for double-stranded DNA cleavage, depending on the substrate sequence. These results have implications for further diversification of the antigen receptor repertoire as well as the role of the RAG proteins in genomic instability.

Base Sequence↗
PubMed2005

IgH V-region sequence does not predict the survival fate of human germinal center B cells.

Germinal center (GC) B cell survival fate is governed in part by the outcome of successful/failed BCR-mediated interactions with accessory cells. However, the extent to which the BCR primary sequence influences such interactions is not fully understood. Over 1000 IgV(H)4 family cDNAs were sequenced from living (annexin V(-)) and apoptotic (annexin V(+) or from within tingible body macrophages) GC B cell fractions from seven tonsils. Results surprisingly demonstrate that living and dying GC B cells do not significantly differ in IgV(H), D, or J(H) gene segment use; HCDR3 length or positive charge; or mutation frequency. Additionally, equivalent IgH cDNA sequences were identified in both fractions, suggesting that BCR sequence alone is an unreliable predictor of GC B cell survival.

Annexin A5↗
PubMed2004

Human fetal, cord blood, and adult lymphocyte progenitors have similar potential for generating B cells with a diverse immunoglobulin repertoire.

Several characteristics of the immunoglobulin (Ig) repertoire in fetuses and adults set them apart from each other. Functionally, this translates into differences in the affinity and effectiveness of the humoral immune response between adults and the very young. At least 2 possibilities could explain these differences: (1) fetal and adult lymphocyte progenitors differ significantly in their potential to form a diverse repertoire, and (2) factors extrinsic to the immunoglobulin locus are more influential to the character of the repertoire. To address this we used nonobese diabetic-severe combined immunodeficient-beta(2) microglobulin knockout (NOD/SCID/beta(2)m(-/-)) mice reconstituted with human B-cell progenitors to compare the immunoglobulin repertoire potential of human fetal, cord blood, and adult sources. We found nearly identical VH and JH gene segment use and only modest differences in the third complementarity determining region of the immunoglobulin heavy chain (HCDR3). We conclude that the repertoire potential is remarkably similar regardless of the age of the individual from which progenitors are derived. Age-related differences in the immunoglobulin repertoire and variance of B-cell responses to immunization appear to arise from selection rather than from changes in recombination of the immunoglobulin locus itself. From the standpoint of the Ig repertoire, an immune system reconstituted from fetal or neonatal stem cells would likely be as diverse as one generated from adult bone marrow.

Adult↗
PubMed2004

Human immunoglobulin selection associated with class switch and possible tolerogenic origins for C delta class-switched B cells.

Current paradigms of peripheral B cell selection suggest that autoreactive B cells are controlled by clonal deletion, anergy, and developmental arrest. We report that changes to the human antibody repertoire likely resulting from these mechanisms both for a well-characterized autoreactivity from antibodies encoded by the V(H)4-34 gene and for other hallmarks of an autoreactive repertoire are apparent mainly for class-switched B cells and not for IgM germinal center, IgM memory, or IgM plasma cells. Other possible indicators of autoreactivity found selected with immunoglobulin class include J(H)6 gene segment usage, increased frequency of B cells with long third hypervariable regions, and distal J(kappa) gene segment bias. Of particular interest is the finding that B cells with these same characteristics are selected into the lineage of B cells that have undergone the unusual class switch from constant region C mu to C delta (C delta-CS). The C delta-CS population also displays an increased frequency of charged amino acids localized to the complementarity-determining regions, further suggesting autoreactivity, and evidence is presented that these B cells had undergone extensive receptor editing. Thus, the C delta-CS lineage may be a "sink" for B cells harboring autoreactive specificities in normal humans. A model for a new tolerizing mechanism that could account for the C delta-CS lineage is presented.

Antibodies, Monoclonal↗
PubMed2004

Ig V region restrictions in human chronic lymphocytic leukemia suggest some cases have a common origin.

The factors that contribute to the development of B cell chronic lymphocytic leukemia (B-CLL) are unknown, and the groups of individuals at the greatest risk for developing this common leukemia are not well defined. Molecular features are important for classifying cases of B-CLL, and it is now apparent that similarities among Ig rearrangements between patients may give important clues to the origin of this disease.

Autoantigens↗
PubMed2003

Alfred Nisonoff.

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Allergy and Immunology↗
PubMed2002

Targeting mucosal sites by polymeric immunoglobulin receptor-directed peptides.

Polymeric immunoglobulins provide first line humoral defense at mucosal surfaces to which they are specifically transported by the polymeric immunoglobulin receptor (pIgR) on mucosal and glandular epithelial cells. Previous studies from our laboratory suggested that amino acids 402-410 of the Calpha3 domain of dimeric IgA (dIgA) represented a potential binding site for the pIgR. Here by binding human secretory component to overlapping decapeptides of Calpha3, we confirm these residues and also uncover an additional site. Furthermore, we show that the Calpha3 motif appears to be sufficient to direct transport of green fluorescent protein through the pIgR-specific cellular transcytosis system. An alternative approach identified phage peptides, selected from a library by the in vitro Madin Darby Canine Kidney transcytosis assay, for pIgR-mediated transport through epithelial cells. Some transcytosis-selected peptides map to the same 402-410 pIgR-binding Calpha3 site. Further in vivo studies document that at least one of these peptides is transported in a rat model measuring hepatic bile transport. In addition to identifying small peptides that are both bound and transported by the pIgR, this study provides evidence that the pIgR-mediated mucosal secretion system may represent a means of targeting small molecule therapeutics and genes to mucosal epithelial cells.

Amino Acid Sequence↗
PubMed2002

Transient association of Ku with nuclear substrates characterized using fluorescence photobleaching.

The autoantigen Ku, composed of subunits Ku70 and Ku86, is necessary for repair of DNA double-strand breaks by nonhomologous end joining. Similarly, Ku participates in repair of DNA double-strand breaks that occur during V(D)J recombination, and it is therefore required for the development of B and T lymphocytes. Although previous studies have identified the DNA-binding activities of Ku, little is known concerning its dynamics, such as the mobility of Ku in the nucleus and its rate of association with substrates. To address this question, fluorescence photobleaching experiments were performed using HeLa cells and B cells expressing a green fluorescent protein (GFP) fusion construct of either Ku70 or Ku86. The results show that Ku moves rapidly throughout the nucleus even following irradiation of the cells. However, the rate of diffusion of Ku was approximately 100-fold slower than that predicted from its size. Association of Ku-GFP with a filamentous nuclear structure was also evident, and nuclear extraction experiments suggest that this represents nuclear matrix. A central domain of Ku70 containing its DNA-binding and heterodimerization regions and its nuclear localization signal shows that this alone is sufficient for the observed mobility of Ku70-GFP and its association with nuclear matrix. These data suggest the mobility of Ku is characterized by a transient, high flux association with nuclear substrates that includes both DNA and the nuclear matrix and may represent a mechanism for repair of double-strand breaks using the nuclear matrix as a scaffold.

Active Transport, Cell Nucleus↗