Novel homozygous silent mutation of ITGB3 gene caused Glanzmann thrombasthenia

Abstract

Glanzmann thrombasthenia (GT) is a rare inherited disease characterized by mucocutaneous bleeding due to the abnormalities in quantity or quality of platelet membrane GP IIb (CD41) or GP IIIa (CD61). GP IIb and GP IIIa are encoded by the ITGA2B and ITGB3 genes, respectively. Herein, we described a 7-year-old Chinese boy of the consanguineous couple who was diagnosed with GT based on the typical clinical manifestations, absence of blood clot retraction and the reduced expression of CD41 and CD61 in platelets. A homozygous silent variant c.1431C > T (p. G477=) of the ITGB3 gene was identified by the Whole-exome sequencing and confirmed by Sanger sequencing. The variant was predicted to affect the splicing. RT-PCR and sequencing revealed that the variant caused a deletion of 95 base pairs and frameshift, and subsequently created a premature stop codon in exon 10 of ITGB3 (p. G477Afs*30). It was indicated that the variant c.1431C > T (p. G477=) of ITGB3 was the cause for Glanzmann thrombasthenia. Our findings expanded the mutation spectrum and provided the information for the genetic counseling, prenatal diagnosis and preimplantation genetic testing (PGT).

Keywords: Glanzmann thrombasthenia; ITGB3; integrin αIIbβ3; silent mutation; whole exome sequencing.

Figure 1

Pedigree of the family and sanger sequencing validation. (A) II₃(proband) had a homozygous mutation (c.1431C > T) in exon 10 of ITGB3 gene. I₁(mother), I₂(father) and II₂(the second elder sister) were all heterozygous carriers. II₁ died at 5 years old and did not carry out any genetic test. (B) Sanger sequencing results of the family members (red arrows indicate the mutation site).

Figure 2

Predict results of the c.1431C > T variant site. (A) The predict result of wild type ITGB3 by using NetGene2 Server (the red square represented). (B) The predict result of ITGB3: c.1431C > T by using NetGene2 Server (the red square represented). (C) The predict result of wild type ITGB3 by using ASSP (the red square represented). (D) The predict result of ITGB3: c.1431C > T by using ASSP (the red square represented).

Figure 3

Analysis of silent variant c.1431C > T (p. G477=) of ITGB3 gene. Sequencing results of ITGB3 cDNA from a healthy control (A) and proband (B). The mutation caused 95 base pairs deletion (c.1430_1524del) and frameshift and then ITGB3 was truncated by a creation of a premature stop codon. (C) Splicing schematic representation of exon 10 organization in wild type of ITGB3 gene. (D) Splicing schematic representation of exon 10 organization in mutant type (ITGB3: c.1431C > T). The amino acid encoded at position 477 was changed from Glycine to Alanine, and caused frameshift. ITGB3 was truncated after 30 amino acids translation (p. G477Afs*30).