Normal polymorphic variations and transcription of the decay accelerating factor gene in paroxysmal nocturnal hemoglobinuria cells.
In paroxysmal nocturnal hemoglobinuria (PNH), an acquired hemolytic anemia, deficiency of decay accelerating factor (DAF) renders blood cells susceptible to increased deposition of autologous complement activation fragments (C3b) and complemented-mediated injury. To investigate the mechanism of the DAF defect, DNA and mRNA from normal and PNH leukocytes were compared in blot hybridization assays by using DAF cDNA and oligonucleotide probes. Southern analyses of DNA from normal cells revealed a single gene spanning approximately equal to 35 kilobases of DNA. Six HindIII banding patterns were distinguishable among normal individuals. In family studies, the patterns segregated as three homozygous and three heterozygous genotypes deriving from three haplotypes: A, B, and C with frequencies of 0.47, 0.36, and 0.17, respectively. Oligonucleotide mapping localized the polymorphic HindIII sites to two noncoding regions in the vicinity of exons encoding (i) the protein oligosaccharide-rich domain and (ii) the mRNA 3'-untranslated region. Analyses of DNA from DAF-negative leukocytes of eight PNH patients demonstrated restriction fragment profiles identical to those of normal individuals for all enzymes studied. Three patients had the BC (normals = 3/32), three patients had the AA (normals = 6/32), and two patients had the AC (normals = 8/32) HindIII genotype. Of the three PNH patients exhibiting the BC genotype, family studies of two demonstrated the expected inheritance patterns, and RNA gel blot analyses of two showed mRNA transcripts indistinguishable from those in normal cells. The absence of DAF gene or mRNA alterations in affected PNH cells that lack other glycolipid-anchored proteins as well as DAF argues that the lesion underlying PNH cells resides in the glycolipid-anchor pathway.
D001798:Blood Proteins / Q000235:genetics*; D018958:CD55 Antigens; D003167:Complement Activation; D005796:Genes; D005819:Genetic Markers; D006457:Hemoglobinuria, Paroxysmal / Q000235:genetics*; D006801:Humans; D007962:Leukocytes / Q000032:analysis; D008565:Membrane Proteins / Q000235:genetics*; D011110:Polymorphism, Genetic; D012150:Polymorphism, Restriction Fragment Length
D001798:Blood Proteins; D018958:CD55 Antigens; D005819:Genetic Markers; D008565:Membrane Proteins