Two New Nonsense Mutations in Type Ia Antithrombin III Deficiency at Leu 140 and Arg 197

Summary

Using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and DNA sequencing, the molecular basis of hereditary type la antithrombin III (AT III) deficiency was disclosed in two families. One mutation was a change from T to A in the codon of TTA for Leu 140 forming a stop codon of TAA, which was confirmed by mutated primer-mediated PCR-HindIII digestion. The application of this method demonstrated that all four affected members had the mutant allele in a heterozygous state and that none of unaffected subjects had this mutation. Another mutation in the second family was a change from C to T in the codon of CGA for Arg 197 also forming a stop codon of TGA, which was confirmed by PCR-HaeIII digestion. Based on these, it was concluded that the two new nonsense mutations in the AT III gene in a heterozygous state are the molecular basis of hereditary type Ia AT III deficiency.

• REFERENCES

• 1 Egeberg O. Inherited antithrombin deficiency causing thrombophilia. Thromb Diath Haemorrh 1965; 13: 516-530
• 2 Cosgriff TM, Bishop DT, Hershgorf EJ, Skolnick MH, Martin BA, Baty BJ, Carlson KS. Familial antithrombin III deficiency: Its natural history, genetics, diagnosis and treatment. Medicine 1983; 62: 209-220
• 3 Lane DA, Ireland H, Olds RJ, Thein SL, Perry DJ, Aiach M. Antithrombin III: A database of mutations. Thromb Haemostas 1991; 66: 657-661
• 4 Bock SC, Wilon KL, Vehar GA, Lawn RM. Cloning and expression of the cDNA for human antithrombin III. Nucl Acids Res 1982; 10: 8113-8125
• 5 Bock SC, Harris JF, Balazs I, Trent JM. Assignment of the human antithrombin III structural gene to chromosome 1q23–25. Cytogenet Cell Genet 1985; 39: 67-69
• 6 Bock SC, Marrinan JA, Radziejewska E. Antithrombin III Utah: Proline-407 to leucine mutation in a highly conserved region near the inhibitor reactive site. Biochemistry 1988; 27: 6171-6178
• 7 Bock SC, Prochownik EV. Molecular genetic survey of 16 kindreds with hereditary antithrombin III deficiency. Blood 1987; 70: 1273-1278
• 8 Olds RJ, Lane DA, Finazzi G, Barbui T, Thein SL. A frameshift mutation leading to type 1 antithrombin deficiency and thrombosis. Blood 1990; 76: 2182-2186
• 9 Olds RJ, Lane DA, Ireland H, Leone G, DeStefano V, Cazenave JP, Wiesel ML, Thein SL. Novel point mutations leading to type Ia antithrombin deficiency and thrombosis. Br J Haematol 1991; 78: 408-413
• 10 Gandrille S, Vidaud D, Emmerich J, Clauser E, Sié P, Fiessinger JN, Alhenc-Gelas M, Priollet P, Aiach M. Molecular basis for hereditary antithrombin III quantitative deficiencies: a stop codon in exon IIIa and a frameshift in exon VI. Br J Haematol 1991; 78: 414-420
• 11 Grundy CB, Thomas F, Millar DS, Krawczak M, Melissari E, Lindo V, Moffat E, Kakkar VV, Cooper DN. Recurrent deletion in the human antithrombin III gene. Blood 1991; 78: 1027-1032
• 12 Vidaud D, Emmerich J, Sirieix ME, Sié P, Alhenc-Gelas M, Aiach M. Molecular basis for antithrombin III type I deficiency: 3 novel mutations located in exon IV Blood. 1991; 78: 2305-2309
• 13 Olds RJ, Lane DA, Ireland H, Finazzi G, Barbui T, Abildgaad U, Girolami A, Thein SL. A common point mutation producing type Ia antithrombin III deficiency. AT 129 CGA to TGA (Arg to stop). Thromb Res 1991; 64: 621-625
• 14 Uno Y, Shigekiyo T, Yoshimoto K, Saito S, Inomoto T. DNA analysis of antithrombin III (AT III) gene in three Japanese kindreds with congenital AT III deficiency. Jpn J Thromb Hemostas 1990; 1: 512-519
• 15 Kusumi S, Kobayashi M, Takata N, Imanaka F, Maehama S, Taketomi Y, Fujimura K, Kuramoto A. Effect of AT III concentrate infusion therapy on the congenital AT III deficiency. Jpn J Clin Hematol 1984; 25: 25-30
• 16 Kunkel LM, Smith KD, Boyer SH, Borgaonkar DS, Wachtel SS, Miller OJ, Breg WR, Jones HW, Rary JM. Analysis of human Y-chromosome-specific reiterated DNA in chromosome variants. Proc Natl Acad Sci USA 1977; 74: 5463-5467
• 17 Iwahana H, Yoshimoto K, Itakura M. Detection of point mutations by SSCP of PCR-amplified DNA after endonuclease digestion. Bio-techniques 1992; 12: 64-66
• 18 Marchuk D, Drumm M, Saulimo A, Collins FS. Construction of T-vectors, a rapid and general system for direct cloning of unmodified PCR products. Nucl Acids Res 1991; 19: 1154
• 19 Schwartz EI, Shevtsov SP, Kuchinski AP, Kovalev YP, Plutalov OV, Berlin YA. Approach to identification of a point mutation in apoB100 gene by means of a PCR-mediated site-directed mutagenesis. Nucl Acids Res 1991; 19: 3752
• 20 Chandra T, Stackhouse R, Kidd VJ, Woo SLC. Isolation and sequence characterization of a cDNA clone of human antithrombin III. Proc Natl Acad Sci USA 1983; 80: 1845-1848
• 21 Winter JH, Bennett B, Watt JL, Brown T, Román CS, Schinzel A, King J, Cook JL. Confirmation of linkage between antithrombin III and Duffy blood group and assignment of AT3 to lq22–25. Ann Hum Genet 1982; 496: 29-34
• 22 Orita M, Suzuki Y, Sekiya T, Hayashi K. Rapid and sensitive detection of point mutations and DNA polymorphisms using the polymerase chain reaction. Genomics 1989; 5: 874-879
• 23 Cooper DN, Krawczak M. Cytosine methylation and the fate of CpG dinucleotides in vertebrate genomes. Hum Genet 1989; 83: 182-188
• 24 Steisinger G, Okada Y, Emrich J, Newton J, Tsugita A, Terzaghi E, Inouye M. Frameshift mutations and the genetic code. Cold Spring Harb Symp Quant Biol 1966; 31: 77-84
• 25 Bock SC, Sibermann JA, Wikoff W, Abildgaard U, Hultin MB. Identification of a threonine for alanine substitution at residue 404 of antithrombin III Oslo suggests integrity of the 404–407 region is important for maintaining normal plasma inhibitor levels. Thromb Haemostas 1989; 62: 494
• 26 Nakagawa M, Tanaka S, Tsuji H, Takada O, Uno M, Hashimoto-Gotoh T, Wagatsuma M. Congenital antithrombin III deficiency (AT-III Kyoto): Identification of a point mutation altering arginine-406 to methionine behind the reactive site. Thromb Res 1991; 64: 101-108
• 27 Miura O, Aoki N. Impaired secretion of mutant α₂-plasmin inhibitor (α2PI-Nara) from COS-7 and HepG2 cells: Molecular and cellular basis for hereditary deficiency of α²-plasmin inhibitor. Blood. 1990; 75: 1092-1096
• 28 Satoh K, Nukiwa T, Brantly M, Garver Jr RI, Hofker M, Curtney M, Crystal RG. Emphysema associated with complete absence of α₁-antitrypsin of a stop codon in an α₁-antitrypsin-coding exon. Am J Hum Genet. 1988; 42: 77-83
• 29 Curiel D, Brantly M, Curiel E, Stier L, Crystal RG. α₁-antitrypsin deficiency caused by the α₁-antitrypsin null_Mattawa gene. An insertion mutation rendering the α₁-antitrypsin gene incapable of producing α₁-antitrypsin. J Clin Invest. 1989; 83: 1144-1152
• 30 Takeshita K, Forget BG, Scarpa A, Benz Jr EJ. Intranuclear defect in β-globin mRNA accumulation due to a premature translation termination codon. Blood 1984; 64: 13-22