Novel RAG1 mutation in a case of severe combined immunodeficiency

Division of Immunology and Allergy, Hospital for Sick Children, University of Toronto, Toronto, Canada.
PEDIATRICS (Impact Factor: 5.3). 10/2005; 116(3):e445-9. DOI: 10.1542/peds.2005-0369
Source: PubMed

ABSTRACT The recombination activating enzymes RAG1 and RAG2 are essential to the process of V(D)J rearrangement in B and T cells and thus to the development of normal immune function. Mutations in RAG1 or RAG2 can lead to a spectrum of disorders, ranging from typical (B-)(T-) severe combined immunodeficiency to Omenn's syndrome. We present a unique presentation of RAG1 deficiency.
We report on a 6-month-old girl who presented with severe respiratory distress, which continued to progress despite antibiotic therapy but seemed to respond to treatment with corticosteroids. The patient exhibited no erythroderma or eosinophilia, and her lymphoid organs were not enlarged.
Investigation of the immune system showed normal numbers of CD3+ T cells, which expressed either CD4 or CD8. Subsequent analysis of the T-cell receptor demonstrated that nearly all CD3+ T cells were clonal; one clone expressed CD4, whereas the other expressed CD8. The extremely restricted T-cell repertoire and the lack of circulating B cells prompted analysis of the RAG1 gene, which revealed a novel homozygous thymine to cytosine substitution at nucleotide position 2686.
This case underscores the importance of more extensive evaluation of the immune system even when widely available, standard, flow cytometric analysis shows normal numbers of T cells that express CD4 or CD8, especially in the absence of circulating B cells.

  • [Show abstract] [Hide abstract]
    ABSTRACT: The recombination-activating gene (RAG) 1/2 proteins play a critical role in the development of T and B cells by initiating the VDJ recombination process that leads to generation of a broad T-cell receptor (TCR) and B-cell receptor repertoire. Pathogenic mutations in the RAG1/2 genes result in various forms of primary immunodeficiency, ranging from T(-)B(-) severe combined immune deficiency to delayed-onset disease with granuloma formation, autoimmunity, or both. It is not clear what contributes to such heterogeneity of phenotypes. We sought to investigate the molecular basis for phenotypic diversity presented in patients with various RAG1 mutations. We have developed a flow cytometry-based assay that allows analysis of RAG recombination activity based on green fluorescent protein expression and have assessed the induction of the Ighc locus rearrangements in mouse Rag1(-/-) pro-B cells reconstituted with wild-type or mutant human RAG1 (hRAG1) using deep sequencing technology. Here we demonstrate correlation between defective recombination activity of hRAG1 mutant proteins and severity of the clinical and immunologic phenotype and provide insights on the molecular mechanisms accounting for such phenotypic diversity. Using a sensitive assay to measure the RAG1 activity level of 79 mutations in a physiologic setting, we demonstrate correlation between recombination activity of RAG1 mutants and the severity of clinical presentation and show that RAG1 mutants can induce specific abnormalities of the VDJ recombination process.
    The Journal of allergy and clinical immunology 11/2013; DOI:10.1016/j.jaci.2013.10.007 · 12.05 Impact Factor
  • Source
    Journal of Allergy and Clinical Immunology 06/2014; 134(4). DOI:10.1016/j.jaci.2014.04.037 · 11.25 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: ζ-associated protein, 70 kd (ZAP70), deficiency in human subjects results in a combined immunodeficiency characterized by normal numbers of circulating CD4 T cells and CD8 lymphocytopenia. Patients who live beyond infancy can also experience autoimmune manifestations. We sought to further characterize the nature of the T-cell populations found in ZAP70-deficient patients and explored the mechanisms that might predispose them to autoimmunity. T-cell development was assessed by examining T-cell receptor (TCR) gene rearrangements and thymopoiesis by measuring TCR exclusion circle levels. TCR repertoire on CD4 and CD8 T-cell populations was assessed by means of flow cytometry. T-cell gene expression patterns were examined by means of exonic microarray analysis and apoptotic responses by means of Annexin V binding and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling. Cells displaying recombination events from all stages of TCR gene rearrangement were present in the peripheral blood of ZAP70-deficient patients; however, the late TCRD-deleting rearrangement was significantly reduced. TCR exclusion circle levels were also found to be low. Surprisingly, all Vβ families were detected in both CD4(+) and CD8(+) circulating T cells. Several Vβ families were significantly overrepresented, which is reminiscent of autoimmune disorders. Levels of mRNA for cytotoxic T lymphocyte-associated antigen 4, TGF-β, and IL-10 were found to be low, a signature of autoimmunity. Finally, Fas-mediated CD4 T-cell apoptosis was found to be reduced in vitro, and staining of thymus biopsy specimens revealed reduced thymocyte apoptosis. We show that in the absence of ZAP70, thymopoiesis is altered and differentiation to double-positive cells is hampered. Circulating T cells appear poorly regulated, do not differentiate into T(H)2 T cells, lack a number of inhibitory growth controls, and display reduced apoptosis, all predisposing patients to exaggerated inflammation and autoimmunity.
    The Journal of allergy and clinical immunology 12/2010; 126(6):1226-33.e1. DOI:10.1016/j.jaci.2010.07.029 · 12.05 Impact Factor