The NF-kappaB canonical pathway is involved in the control of the exonucleolytic processing of coding ends during V(D)J recombination.
V(D)J recombination is essential to produce an Ig repertoire with a large range of Ag specificities. Although NF-kappaB-binding sites are present in the human and mouse IgH, Igkappa, and Iglambda enhancer modules and RAG expression is controlled by NF-kappaB, it is not known whether NF-kappaB regulates V(D)J recombination mechanisms after RAG-mediated dsDNA breaks. To clarify the involvement of NF-kappaB in human V(D)J recombination, we amplified Ig gene rearrangements from individual peripheral B cells of patients with X-linked anhidrotic ectodermal dysplasia with hyper-IgM syndrome (HED-ID) who have deficient expression of the NF-kappaB essential modulator (NEMO/Ikkgamma). The amplification of nonproductive Ig gene rearrangements from HED-ID B cells reflects the influence of the Ikkgamma-mediated canonical NF-kappaB pathway on specific molecular mechanisms involved in V(D)J recombination. We found that the CDR3(H) from HED-ID B cells were abnormally long, as a result of a marked reduction in the exonuclease activity on the V, D, and J germline coding ends, whereas random N-nucleotide addition and palindromic overhangs (P nucleotides) were comparable to controls. This suggests that an intact canonical NF-kappaB pathway is essential for normal exonucleolytic activity during human V(D)J recombination, whereas terminal deoxynucleotide transferase, Artemis, and DNA-dependent protein kinase catalytic subunit activity are not affected. The generation of memory B cells and somatic hypermutation were markedly deficient confirming a role for NF-kappaB in these events of B cell maturation. However, selection of the primary B cell repertoire appeared to be intact and was partially able to correct the defects generated by abnormal V(D)J recombination.
Souto-Carneiro
M Margarida
MM
Autoimmunity Branch, National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD 20892-1560, USA.
Fritsch
Ruth
R
Sepúlveda
Nuno
N
Lagareiro
M João
MJ
Morgado
Nuno
N
Longo
Nancy S
NS
Lipsky
Peter E
PE
D000595:Amino Acid Sequence; D001402:B-Lymphocytes / Q000276:immunology; D053358:Ectodermal Dysplasia 1, Anhidrotic / Q000235:genetics*; D005092:Exonucleases / Q000378:metabolism; D015322:Gene Rearrangement, B-Lymphocyte; D015326:Gene Rearrangement, B-Lymphocyte, Heavy Chain; D015328:Gene Rearrangement, B-Lymphocyte, Light Chain; D006801:Humans; D053306:Hyper-IgM Immunodeficiency Syndrome / Q000235:genetics*; D007072:Immunoglobulin D / Q000032:analysis; D008297:Male; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7 / Q000032:analysis
D007072:Immunoglobulin D; D016328:NF-kappa B; D018127:Tumor Necrosis Factor Receptor Superfamily, Member 7; D005092:Exonucleases