deoxyguanosine-triphosphate and Purine-Pyrimidine-Metabolism--Inborn-Errors

Topics: Deoxyguanine Nucleotides; Diagnosis, Differential; Humans; Immune System Diseases; Mutation; Prognosis; Purine-Nucleoside Phosphorylase; Purine-Pyrimidine Metabolism, Inborn Errors; T-Lymphocytes; Uric Acid

The genetic deficiencies of adenosine deaminase and purine nucleoside phosphorylase lead to blocks in the purine pathway. The intracellular accumulation of deoxynucleosides and deoxynucleotides is toxic to both dividing and nondividing lymphocytes via multiple mechanisms. T-lymphocytes are uniquely sensitive to purine-mediated cytotoxicity because of a functional imbalance of phosphorylating and dephosphorylating enzymatic activities. These inborn errors or purine metabolism are rare disorders. The study of these conditions, however, has uncovered unique enzymatic properties of lymphocytes and lymphocyte subclasses. A better understanding of the mechanisms of lymphocytotoxicity in these two purine enzyme defects may lead to better modes of therapeutic manipulation of the immune system.

Topics: Adenosine Deaminase; Cytotoxicity, Immunologic; Deoxyadenine Nucleotides; Deoxyadenosines; Deoxyguanine Nucleotides; Humans; Immunologic Deficiency Syndromes; Lymphocytes; Purine-Nucleoside Phosphorylase; Purine-Pyrimidine Metabolism, Inborn Errors; Purines

Template-independent nucleotide additions (N regions) generated at sites of V(D)J recombination by terminal deoxynucleotidyl transferase (TdT) increase the diversity of antigen receptors. Two inborn errors of purine metabolism, deficiencies of adenosine deaminase (ADA) and purine nucleoside phosphorylase (PNP), result in defective lymphoid development and aberrant pools of 2'-deoxynucleotides that are substrates for TdT in lymphoid precursors. We have asked whether selective increases in dATP or dGTP pools result in altered N regions in an extrachromosomal substrate transfected into T-cell or pre-B-cell lines. Exposure of the transfected cells to 2'-deoxyadenosine and an ADA inhibitor increased the dATP pool and resulted in a marked increase in A-T insertions at recombination junctions, with an overall decreased frequency of V(D)J recombination. Sequence analysis of VH-DH-JH junctions from the IgM locus in B-cell lines from ADA-deficient patients demonstrated an increase in A-T insertions equivalent to that found in the transfected cells. In contrast, elevation of dGTP pools, as would occur in PNP deficiency, did not alter the already rich G-C content of N regions. We conclude that the frequency of V(D)J recombination and the composition of N-insertions are influenced by increases in dATP levels, potentially leading to alterations in antigen receptors and aberrant lymphoid development. Alterations in N-region insertions may contribute to the B-cell dysfunction associated with ADA deficiency.

Topics: Adenosine Deaminase; Adenosine Deaminase Inhibitors; Adenosine Triphosphate; Cells, Cultured; Deoxyadenosines; Deoxyguanine Nucleotides; Deoxyribonucleotides; DNA Nucleotidylexotransferase; Gene Rearrangement, B-Lymphocyte; Humans; Immunoglobulin M; Purine-Pyrimidine Metabolism, Inborn Errors; Recombination, Genetic; Sequence Analysis, DNA

Using HPLC methods, we measured the concentrations of nucleosides and nucleotides for a patient with no purine nucleoside phosphorylase (PNP; EC 2.4.2.1) enzymatic activity. Concentrations of inosine and guanosine were abnormally high in urine and plasma, whereas guanosine diphosphate (GDP) and guanosine triphosphate (GTP) concentrations in erythrocytes were depleted. The unusual presence of deoxyribonucleosides (deoxyinosine and deoxyguanosine) and deoxyribonucleotides (dGDP and dGTP) was also notable. Thus, HPLC represents an accurate and useful tool for the study of purine metabolic disorders.

Topics: Chromatography, High Pressure Liquid; Deoxyadenine Nucleotides; Deoxyguanine Nucleotides; Erythrocytes; Guanosine; Guanosine Diphosphate; Guanosine Triphosphate; Humans; Infant; Inosine; Male; Purine-Nucleoside Phosphorylase; Purine-Pyrimidine Metabolism, Inborn Errors