concanavalin-a and Severe-Combined-Immunodeficiency

The interleukin-2 (IL-2) receptor gamma chain is indispensable for IL-2-, IL-4-, IL-7-, IL-9-, and IL-15-mediated signaling. Mutations of the human gamma chain cause the X-linked severe combined immunodeficiency (XSCID), showing that T and natural killer cells absolutely require the gamma chain for their development in humans. To elucidate the roles of the gamma chain in hematopoiesis, we have generated mice, by gene targeting, that express a form of the gamma chain lacking the cytoplasmic region. Male mice carrying the truncated gamma-chain mutant, which mimics mutations in patients with XSCID, showed a decrease in the number of lymphocytes and an increase in monocytes; the number of T cells was profoundly reduced and no natural killer cells were detected, which is similar to the characteristic of human XSCID. Unlike human XSCID, the levels of B cells were also reduced. In spite of the severe decrease in CD45R+/sIgM+ B cells, the level of IgM in serum of the 8-week-old mutant mice was higher than that of control littermates. Interestingly, the stem cell population with surface phenotypes of CD34, c-kit, and Sca-1 was significantly increased. Furthermore, the colony-forming assay showed that the mutant mice had 15-fold higher numbers of hematopoietic progenitor cells in the spleen as compared with that of controls. These results indicate that functional loss of the gamma chain causes significant effects on the immunological system in mice.

Topics: Animals; B-Lymphocyte Subsets; Bone Marrow; Concanavalin A; Disease Models, Animal; Gene Expression; Gene Targeting; Hematopoiesis; Hematopoietic Stem Cells; Humans; Immunoglobulin M; Immunophenotyping; Lymphocyte Activation; Lymphocyte Count; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Receptors, Interleukin-2; Sequence Deletion; Severe Combined Immunodeficiency; Signal Transduction; Spleen; T-Lymphocyte Subsets; Thymus Gland; X Chromosome

Mutations at the adenosine deaminase (ADA) locus result in a spectrum of disorders, encompassing a fulminant neonatal onset severe combined immunodeficiency (SCID) and childhood onset immunodeficiency, as well as apparently normal immune function. The extent of accumulation of the toxic metabolite, deoxyATP, correlates directly with severity of disease. We have now determined the mutations on both alleles of a child with fulminant, neonatal onset ADA- SCID and accumulation of extremely high concentrations of deoxyATP. The genotype was consistent with the severely affected phenotype. One allele carried a large deletion that arose by non-homologous recombination and included the first five exons and promoter region. The second allele carried a missense mutation (G649A) resulting in replacement of Glu217, an amino acid involved in the catalytic site, by Lys and predicting a major alteration in charge. Expression of the mutant cDNA in Cos cells confirmed that the mutation abolished enzyme activity. We have previously reported that a missense mutation at the preceding codon is similarly associated with neonatal onset ADA- SCID and accumulation of extremely high deoxyATP. These findings suggest that genotype-phenotype correlations may be apparent for ADA- SCID, despite the role that random variation in exposure to environmental pathogens may play in the initial phenotype. Such genotype-phenotype correlations may be important to consider in evaluating results of ongoing trials of "gene" and enzyme replacement therapy.

Topics: Adenosine Deaminase; Alleles; Ampicillin; Base Sequence; Chromosomes, Human, Pair 2; Concanavalin A; Deoxyadenine Nucleotides; Deoxyadenosines; DNA Probes; Drug Therapy, Combination; Gene Library; Gentamicins; Humans; Infant; Ketoconazole; Lymphocyte Activation; Male; Molecular Sequence Data; Oxacillin; Phytohemagglutinins; Point Mutation; Pokeweed Mitogens; Promoter Regions, Genetic; Restriction Mapping; Sequence Homology, Nucleic Acid; Severe Combined Immunodeficiency; Trimethoprim, Sulfamethoxazole Drug Combination