A three amino acid deletion in glycoprotein IIIa is responsible for type I Glanzmann's thrombasthenia: importance of residues Ile325Pro326Gly327 for beta3 integrin subunit association
| Autor: | Mc, Morel-Kopp, Kaplan C, valerie proulle, Jallu V, Melchior C, Peyruchaud O, Mh, Aurousseau, Kieffer N |
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| Rok vydání: | 1997 |
| Předmět: |
Male
Proline Glycine CHO Cells Platelet Membrane Glycoproteins Transfection Polymerase Chain Reaction Consanguinity Antigens CD Cricetinae Animals Humans Amino Acid Sequence Isoleucine Polymorphism Single-Stranded Conformational DNA Primers Sequence Deletion Base Sequence Integrin beta3 Infant Recombinant Proteins Pedigree Algeria Female France Thrombasthenia |
| Zdroj: | Europe PubMed Central |
| ISSN: | 0006-4971 |
| Popis: | Glanzmann's thrombasthenia (GT) is a recessive autosomal bleeding disorder characterized by abnormal platelet aggregation due to a qualitative or quantitative defect of the glycoprotein (GP) IIb-IIIa complex (integrin alphaIIb beta3). We describe a new mutation in the GPIIIa gene responsible for type I GT in a consanguineous Algerian family. A discordance between phenotyping and genotyping of the GPIIIa-related HPA-1 platelet alloantigen system in three family members heterozygous for the disease suggested a genetic defect in the GPIIIa gene and a normal GPIIb gene. Sequence analysis of amplified genomic DNA fragments showed a 6-bp deletion in exon 7 of the GPIIIa gene resulting in the amino acid deletion/substitution (Ile325pro326Gly327 --Met) and creating a new BspHI restriction site. Expression of the mutated integrin beta3 subunit cDNA in Chinese hamster ovary cells showed that the cDNA gene was transcribed into a full-length beta3 protein with an apparent molecular weight identical to wild-type beta3 and accumulated as a single-chain molecule in the cell cytoplasm. The absence of heterodimeric complex formation of the mutant beta3 protein with endogenous alpha v was shown by immunoprecipitation experiments, intracellular immunofluorescent labeling, and a semiquantitative enzyme-linked immunosorbent assay using the alpha vbeta3 complex-specific monoclonal antibodies LM609 and 23C6. Substitution of the methionine residue by a proline, present at position 326 of wild-type beta3, did not restore the ability of the recombinant mutant beta3 protein to associate with alpha v , suggesting that the Ile-Pro-Gly motif is located in a beta3 domain important for integrin subunit interaction. The association of a BspHI restriction site with this newly identified mutation has allowed allele-specific restriction analysis of Algerian GT individuals and the identification of two new unrelated type I patients exhibiting the same mutation, suggesting that the described mutation might be significant in this population and that BspHI restriction analysis will provide a useful screening assay for antenatal diagnosis and genetic counselling. |
| Databáze: | OpenAIRE |
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